The Placenta: Basics and Clinical Significance 3662662558, 9783662662557

Table of contents :
Foreword
Contents
Editors and Contributors
About the Editors
Contributors
Abbreviations
1: Placental Development with Histological Aspects
1.1 Introduction
1.2 Development of the Placenta
1.2.1 Prelacunar Phase (Days 5 to 8 after fertilization)
1.2.2 Lacunar Phase (Days 8 to 13 after fertilization)
1.2.3 Early Villous Phase (Days 13 to 28 p. c.)
1.2.4 Villous Phase (Day 28 p. c. to end of pregnancy)
1.3 Placental Villi
1.3.1 General Histological Structure
1.3.2 Syncytiotrophoblast
In Vitro Culture and Syncytiotrophoblast
Structure and Function of the Syncytiotrophoblast
Protrusions of the Syncytiotrophoblast
1.3.3 Villous Cytotrophoblast (Langhans Cell)
1.3.4 Villous Stroma
1.3.5 Placental Blood Vessels
1.4 Architecture of the Villous Tree
1.4.1 Mesenchymal Villi
1.4.2 Immature Intermediate Villi
1.4.3 Stem Villi
1.4.4 Mature Intermediate Villi
1.4.5 Terminal Villi
1.5 Extravillous Trophoblast
1.5.1 Interstitial Trophoblast
1.5.2 Endoglandular Trophoblast
1.5.3 Endovascular Trophoblast
Endoarterial Trophoblast
Endovenous Trophoblast
1.5.4 Endolymphatic Trophoblast
1.5.5 General Considerations on Trophoblast Invasion
1.5.6 Maternal Perfusion of the Placenta
In the First Trimester
In the Second and Third Trimester
1.5.7 Formation of the Chorion laeve (fetal membranes)
References
2: Immunology of the Fetomaternal Border
2.1 Background
2.2 Immunology of the Endometrium
2.3 Immunology of Pregnancy
2.3.1 Problems
2.3.2 Fetomaternal Interfaces
2.3.3 Pregnancy: A Th2 Phenomenon
2.3.4 Regulatory T Cells
2.3.5 γδ-T Cells
2.3.6 Natural Killer Cells
Uterine NK Cells
2.3.7 Uterine CD14+ Cells
2.3.8 Trophoblastic Immunoregulatory Factors
HLA Class Ib
Indoleamine 2,3-dioxygenase (IDO)
2.3.9 Hormones
Human Chorionic Gonadotropin (hCG)
2.3.10 Other Selected Pregnancy-promoting Mechanisms
Fas/Fas Ligand
Galectin-1
Trophoblastic Extracellular Vesicles
2.4 Summary and Conclusion
References
3: Placental Morphology
3.1 Morphology of the Placenta
3.1.1 Introduction
3.1.2 Villous Development
3.1.3 Development of Cell Columns for Trophoblast Invasion
3.1.4 Structures on the Villous Surface
3.2 Histopathology of the Placenta for Gynecologists
3.2.1 Introduction
3.2.2 First Trimester (Abortion)
Abortion with and Without Developmental Aberration
Question About Hydatidiform Mole
Answers and Evidence from Examination of Abortion Material
3.2.3 Second Trimester (Hydrops Fetalis, Infection and Inflammation)
Hydrops of Fetus and Placenta
Immunological Hydrops in Blood Group Incompatibility
Hydrops as A Result of Infection (Fifth Disease, Erythema infectiosum)
Placentitis
(Acute) Inflammation of the Chorionic Villi with Evidence of Pathogens
(Chronic) Inflammation of the Chorionic Villi Without Evidence of Pathogens
Chorioamnionitis (Amniotic Infection Syndrome)
Umbilical Cord Infection
Answers and Evidence from Examination of Placentas in Preterm Delivery
3.2.4 Third Trimester (Circulatory and Maturation Disorders)
Maternal Circulatory Disorder
Preeclampsia
Premature Placental Abruption
Fetal Circulatory Disorder
Endangiopathy Obliterans
Hereditary Thrombophilias
Umbilical Cord Complications
Maturation Disorder
Intermediate Villus Deficiency
Chorangiosis
Maturation Retardation
Terminal Villus Deficiency
Responses and Evidence from Studies of Small-for-date Newborns and Stillbirths
3.2.5 Postpartum Period
Placenta Accreta or Increta
Neoplasia
Malignant Degeneration of the Chorial Tissue
Metastases to the Placenta
Twins
Chorionicity and Zygosity
Fetofetal Transfusion
3.3 Biobanking
3.3.1 Introduction
What Exactly Is Pre-analysis?
3.3.2 Variables Affecting the Composition of a Sample
Variables Before Sampling
Variables After Sampling
3.3.3 Collection or Biobank?
Definition of Biobanks
Advantages of Biobanks
Further Readings
Further Reading on Section 3.2
4: Placental Function—Nutrient Transport—Gas Exchange
4.1 General Functions of the Placenta
4.2 Nutrient Transport Across the Placenta
4.2.1 Transport of Lipids and Fatty Acids
4.2.2 Transport of Glucose
4.2.3 Transport of Proteins and Amino Acids
4.2.4 Transport of Minerals and Trace Elements
4.3 Maternofetal Gas Exchange
References
5: Endocrinology of the Placenta
5.1 Introduction
5.2 Steroid Hormones
5.2.1 Progesterone
5.2.2 Estrogens
5.2.3 Glucocorticoids
5.3 Peptide Hormones
5.3.1 Human Chorionic Gonadotropin (hCG)
5.3.2 Leptin
5.3.3 Corticotrophin Releasing Hormone (CRH)
5.3.4 Placental Lactogen (hPL) and Placental Growth Hormone (hPGH)
5.3.5 Summary
References
6: Teratology
6.1 Introduction
6.2 Congenital Anomalies Historically
6.3 Congenital Anomalies Today
6.4 Basic Risk
6.5 Medication and Pregnancy
6.6 Placenta and Teratology
6.6.1 The Sensitivity of the Embryo to Toxic Influences Depends on the Genotype
6.6.2 The Sensitivity of the Embryo to Toxic Influences Depends on Its Stage of Development
6.6.3 Different Embryotoxic Influences Affect (Embryonic) Development via Relatively Few Specific Mechanisms
6.6.4 After Exposure to Teratogens, Different Developmental Courses Are Possible in Principle
6.6.5 The Way in Which Toxic Influences Reach the Embryo/Fetus Depends on Their Physical and Chemical Properties
6.6.6 Dose-response Relationships Apply in Teratology as Elsewhere in Pharmacology and Toxicology
6.7 Conclusion
References
7: The Effects of Legal and Illegal Drugs on Placental Function
7.1 Introduction
7.2 Smoking During Pregnancy
7.2.1 Tobacco Ingredients
7.2.2 Effects on Placental Morphology
7.2.3 Effects on Trophoblast Cells
7.2.4 Oxidative Stress and Endothelial Dysfunction
7.2.5 Placental Transcriptome
7.3 Alcohol During Pregnancy
7.3.1 Effects on Placental Morphology
7.3.2 Effects on Trophoblast Cells
7.3.3 Oxidative Stress and Endothelial Dysfunction
7.4 Methamphetamines and MDMA
7.5 Cocaine
7.6 Opiates
7.6.1 Placental Transfer
7.6.2 Effects on Trophoblast Cells
7.7 Cannabis
7.7.1 Placental Transfer
7.7.2 Effects on Trophoblast Cells
References
8: Placenta-Related Hemorrhage: Pathophysiology, Diagnostics, Management
8.1 The Placenta Accreta Spectrum (PAS)
8.1.1 Introduction
8.1.2 Epidemiology
8.1.3 Risk Factors
8.1.4 Definition
8.1.5 Pathogenesis
8.1.6 Diagnosis
Ultrasound for the Diagnosis of PAS
Thinning or Absence of the Myometrium
Placental Protrusion
Focal Expophytic Excrescence
Loss of the Clear Zone
Abnormal Placental Lacunae
Urinary Bladder Disruption
Uterovesical Hypervascularity
Subplacental Hypervascularity
Bridging Vessels
Vessels Feeding the Placental Lacunae: Lacunae Feeder Vessels
Summary Ultrasound
MRI for the Diagnosis of PAS
Laboratory Tests
8.1.7 Management and Therapy
Possible Procedure in case of Antenatal Diagnosis
Antepartum Management
Surgical Procedures
Conservative Procedure
Possible Procedure in case of Intrapartum Diagnosis
8.2 Placenta Praevia
8.2.1 Terminology
8.2.2 Morbidity and Mortality
8.2.3 Etiology and Risk Factors
Special Case Placenta Praevia Accreta
8.2.4 Diagnostics and Management
Clinical Management
8.2.5 Operational Procedure
Emergency Situation
Placenta Praevia, Originating from the Posterior Wall
Placenta Praevia, Originating from the Anterior Wall
Atony Risk
8.2.6 Summary
8.3 Umbilical Cord Insertion, Variations and Vasa Praevia
8.3.1 Umbilical Cord Insertion, Velamentous Cord Insertion
8.3.2 Vasa Praevia
8.4 Premature Placental Abruption
8.4.1 Incidence and Risk Factors
8.4.2 Definition
8.4.3 Etiology
8.4.4 Clinical Signs
8.4.5 Instrumental Diagnostics
B-Mode Sonography
Doppler Sonography
Cardiotocography (CTG)
Magnetic Resonance Imaging (MRI) and Computed Tomography (CT)
8.4.6 Laboratory Diagnostics
Serum Markers in the First and Second Trimester
Kleihauer-Betke Test
Blood Count/Coagulation Diagnostics
8.4.7 Clinical Care/Management
8.4.8 Conclusion
8.5 Primary and Secondary Tumors of the Umbilical Cord and Placenta
8.5.1 Tumors of the Umbilical Cord
8.5.2 Tumors of the Placenta
Primary Non-trophoblastic Tumors of the Placenta
Teratomas
Chorangiomas, Fibromas and Chorangiocarcinoma
Leiomyomas, Endometrial Stromal Tumors, Deciduomas, Hepatocellular Adenomas
Intraplacental Heterotopias
Secondary Tumors of the Placenta
Maternal Tumors
Fetal Tumors
Primary Trophoblastic Tumors of the Placenta
References
References on Section 8.1
References on Section 8.2
References on Section 8.3
References on Section 8.4
References on Section 8.5
9: Placental Imaging
9.1 Sonographic Assessment of the Placenta in the Second and Third Trimester and Ultrasound/MRI Morphology of the Placenta
9.1.1 Introduction
9.1.2 Localization
Low-Lying Placenta
Placenta Praevia
Untypical Placental Location
9.1.3 Placenta Accreta Spectrum (PAS)
Imaging
9.1.4 Echogenicity
Placental Lacunae
Septal Decidual Cysts
Placental Cysts of the Surface
Echogenic Cystic Lesions (Placental Bed Infarction)
Placental Infarction
Placental Hematoma
Breus’ Mole
Jelly-like Placenta
Mesenchymal Dysplasia of the Placenta
9.1.5 Maturation of the Placenta
9.1.6 Size and Shape
Accessory Placenta (Placenta succenturiata)
Placenta circumvallata
Placenta membranacea/Placenta diffusa
9.1.7 Placental Biometry and Volumetry
Thick Placenta
Small Sized Placenta
Placental Abruption
9.1.8 Summary
9.2 Doppler Sonography/Functional Diagnostics
9.2.1 Placental Vascular System
Fetoplacental Hemodynamics
Uteroplacental Hemodynamics
Disorders of Utero- and Fetoplacental Blood Flow
Placental Function
Placental Blood Flow
Oxygenation and Placental Metabolism
References
Section 9.1
Section 9.2
10: Disorders of Early Pregnancy and Pregnancy Loss
10.1 Early Pregnancy and Its Disturbance
10.1.1 Diagnosis of Early Pregnancy
General Remarks
Laboratory Diagnostics
Human Chorionic Gonadotropin (hCG)
Additional Factors, Hormones and Screening Tests
Verification of Early Pregnancy by Means of Sonography
10.1.2 Pregnancy Loss
Definition and Epidemiology
Clinical Stages of the Abortion Process
Causes of Pregnancy Loss
Chromosomal Abnormalities in Pregnancy Tissue
Genetic Causes in the Parents
Uterine Anomalies
Congenital Uterine Anomalies
Acquired Uterine Anomalies
Infections
Stimulants and Pollutants
Endocrine Causes
Psychosocial Factors
10.2 Recurrent Pregnancy Loss
10.2.1 Introduction
10.2.2 Established Risk Factors
Endocrine Dysfunctions
Anatomical Malformations
Infections
Chromosomal Disorders
Psychological Factors
Hemostasiological Factors
Immunological Factors
10.2.3 Possible New Risk Factors
Chronic Endometritis
Peripheral and Uterine Killer Cells
References
11: Placental Insufficiency/Placenta-Associated Diseases
11.1 Placental Disorders: Pathophysiology
11.1.1 Introduction
11.1.2 Preeclampsia
Definition
Classification and Epidemiology
Long-Term Effects
Pathophysiology
Risk Factors for the Development of Preeclampsia
Differences Between Early and Late Onset Preeclampsia
Presentation and Falsification of the Outdated Hypothesis on the Etiology of Preeclampsia
Deficient Trophoblast Invasion and Preeclampsia
Deficient Placental Perfusion, Placental Hypoxia and Preeclampsia
What Is the Effect of the Funnel-Shaped Dilatation of the Uterine Arteries?
What Is the Effect of Deficient Trophoblast Invasion?
Does This Really Lead to Deficient Placental Perfusion and Placental Hypoxia?
Possible Explanation of the Etiology of Preeclampsia
11.2 FGR: Diagnostics and Management
11.2.1 Terminology and Definition
11.2.2 Epidemiology
11.2.3 Cause and Risk Factors
11.2.4 Fetal Compensation
11.2.5 Outcome of the Child
11.2.6 Diagnostics and Monitoring
Growth Dynamics
Head-Abdomen Discrepancy
Amniotic Fluid Volume
Biophysical Profile
Doppler Ultrasound
Umbilical Arteries and Descending Aorta
Middle Cerebral Artery and Cerebroplacental Ratio (CPR)
Ductus Venosus
11.2.7 Therapeutic Management–Delivery Indication
Growth Restriction Intervention Trial (GRIT)
Disproportionate Intrauterine Growth Intervention Trial at Term (DIGITAT)
Trial of Randomized Umbilical and Fetal Flow in Europe (TRUFFLE)
11.3 Preeclampsia: Diagnosis and Management
11.3.1 Definition and Classification
11.3.2 Causes and Risk Factors
11.3.3 Diagnosis and Early Detection
11.3.4 Biomarkers in Diagnostics and Prediction
11.3.5 Risk Assessment for Preeclampsia in the Context of First Trimester Screening and Secondary Prophylaxis
11.3.6 Clinical Management
11.3.7 Possible Future Therapeutic Approaches
11.3.8 Long-Term Morbidity
References
Section 11.1
Section 11.2
Section 11.3
12: The Placenta in Twins
12.1 Introduction
12.2 Structural Differences Between Monochorial and Dichorial Placentas
12.2.1 Chorionicity
12.2.2 Assessment of the Placenta in Early Pregnancy
12.2.3 Assessment of the Placenta in Late Pregnancy
12.2.4 Assessment of the Placenta after Delivery
12.3 The Placenta in Twin-to-Twin Transfusion Syndrome (TTTS)
12.3.1 Vascular Anastomoses in TTTS
12.3.2 Importance of the Umbilical Cord in TTTS
12.3.3 Unequal Division of the Placenta in TTTS
12.3.4 Significance of Other Placental Factors for TTTS
12.4 Monochorial Placenta and Discordant Growth
12.4.1 Unequal Division of the Placenta and Discordant Growth
12.4.2 Anastomoses and Discordant Growth
12.4.3 Umbilical Cord and Growth Discordance
12.4.4 Molecular Changes and Discordant Growth
12.4.5 Other Placental Factors and Discordant Growth
12.5 Dichorial Placenta and Discordant Growth
12.5.1 Umbilical Cord and Discordant Growth
12.5.2 Placental Pathology and Discordant Growth
12.6 Conclusion
References
13: Fetal Programming
13.1 Introduction
13.2 Between Hypothesis and Epidemiology
13.2.1 This Is How It All Began: The Barker Hypothesis and First Epidemiological Studies
13.2.2 Obesity, Insulin Resistance and Metabolic Syndrome
13.2.3 Cardiovascular Diseases
13.2.4 Altered Immune Response and Autoimmune Diseases
13.2.5 Memory and Psychiatric Disorders
13.2.6 Gender-specific Programming
13.3 Underlying Mechanisms
13.3.1 Direct Mother-to-Child Mediators
Maternal Hormones, Growth Factors and Cytokines
Hypothalamic-Pituitary-Adrenal (HPA) Axis
Signaling Pathways Via Placentally-expressed Growth Factors
Maternal Microchimerism
13.3.2 Epigenetic Changes
13.4 Examples of Exogenous Stimuli of Fetal Programming
13.4.1 Maternal Malnutrition and Placental Insufficiency
13.4.2 Maternal Oversupply and Gestational Diabetes
13.4.3 Glucocorticoid Administration for Lung Maturation Induction
13.5 Contradictions and Alternative Approaches
13.6 Pregnancy as an Option for Future Health Prevention
References
14: Fetal Cells and Cell-Free Nucleic Acids in Maternal Blood: Genetic and Immunological Aspects
14.1 Introduction
14.2 Fetal Cells and Fetal DNA in Maternal Blood: A Centuries-Old Phenomenon
14.3 Cell-Free DNA: Current Status for Non-invasive Prenatal Testing
14.3.1 Whole Genome Sequencing
14.3.2 Targeted Genome Sequencing
14.3.3 SNP Approach
14.3.4 Technical Principles of Currently Available Non-invasive Prenatal Tests
14.3.5 Clinical Use of NIPT, Its Limitations and Impact on Prenatal Care
14.4 Trophoblast Cells in the Cervix – Diagnostic Use and Insight into Fetomaternal Pathologies
14.5 Placental Cell-Free DNA: An Activator of the Maternal Immune System and Initiator of Birth?
14.6 MicroRNA from the Placenta: New Antiviral Agents?
14.7 Circulating Fetal Cells: Their Role in Pregnancy-Associated and Postpartum Diseases
14.8 Conclusions
References
15: Research Aspects and In Vitro Models
15.1 Trophoblast Cell Culture Models
15.1.1 Primary Trophoblasts
15.1.2 Trophoblast Cell Lines
Choriocarcinoma Cell Lines
Trophoblast Hybridoma Cells
Transformed Trophoblasts
15.2 Placental Explant Cultures
15.3 Endothelial Cells
15.3.1 Human Umbilical Vein Endothelial Cells (HUVECs)
15.3.2 Primary Placental Endothelial Cells
15.4 Placental Macrophages
15.4.1 Decidual Macrophages
15.4.2 Hofbauer Cells
15.5 Placenta Ex Vivo Perfusion
15.5.1 Methodology
15.5.2 Applications in Research
References
16: Maternal Disease Affecting the Placenta: Diabetes Mellitus
16.1 Definition and Epidemiology
16.1.1 Definition
16.1.2 Epidemiology
16.2 Effects of Hyperglycemia in Pregnancy
16.3 Diagnosis and Therapy
16.3.1 Screening and Diagnosis of Gestational Diabetes
16.3.2 Therapy
16.3.3 Timing of Delivery in Patients with Diabetes
16.4 Significance of the Placenta for Glucose Metabolism in Pregnancy
16.5 Placental Changes in Patients with Diabetes
16.5.1 Placental Changes in Pre-Existing Diabetes
16.5.2 Placental Changes in Gestational Diabetes
16.6 Placental Histology in Diabetes—Consequences of a Histopathological Reprocessing?
References
Index

Citation preview

The Placenta Basics and Clinical Significance Berthold Huppertz Ekkehard Schleußner Editors

123

The Placenta

Berthold Huppertz • Ekkehard Schleußner Editors

The Placenta Basics and Clinical Significance

Editors

Berthold Huppertz Division of Cell Biology Histology and Embryology Gottfried Schatz Research Center Medical University of Graz Graz, Austria

Ekkehard Schleußner Clinic for Gynecology and Obstetrics University Hospital Jena Jena, Germany

ISBN 978-3-662-66255-7 ISBN 978-3-662-66256-4 (eBook) https://doi.org/10.1007/978-3-662-66256-4 © Springer-Verlag GmbH Germany, part of Springer Nature 2023 The translation was done with the help of artificial intelligence (machine translation by the service DeepL. com). A subsequent human revision was done primarily in terms of content. This work is subject to copyright. All rights are reserved by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed. The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use. The publisher, the authors, and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty, expressed or implied, with respect to the material contained herein or for any errors or omissions that may have been made. The publisher remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. This Springer imprint is published by the registered company Springer-Verlag GmbH, DE, part of Springer Nature. The registered company address is: Heidelberger Platz 3, 14197 Berlin, Germany

V

Foreword Dear Reader, Why a textbook on the placenta? Every person has this organ for only 9 months, after which it is disposed of or, at best, a tree is planted on it. And yet it is one of the most exciting subjects imaginable! Within a very short time, the placenta has to develop, take up its function, make a successful pregnancy possible, supply and protect the unborn child—and at the same time influence all bodily functions of the pregnant woman in such a way that the resources are also available for all the above-mentioned processes. When Springer-Verlag asked whether I, Berthold Huppertz, would like to publish a textbook on the placenta, the publishing staff did not know that this was and is a long-cherished wish of me as an enthusiastic placentologist. Now there is the challenge of publishing a textbook on the placenta that would meet my requirements as a basic scientist, but at the same time be appealing to clinically active colleagues. It was therefore clear at a very early stage that an obstetrician had to be found as a partner. This was quickly found in Ekkehard Schleußner—with his “favourite organ, the placenta.” Together we defined topics, found friends and colleagues as co-authors, and finally we dared to tackle this project. There are many textbooks in the fields of pregnancy and obstetrics, and some of them have small sections on the placenta. Some decades ago, there was also a very good atlas of placental morphology by Martin Vogel—also published by Springer. But until today (until this book!), there is no textbook on the human placenta in German language, which covers and combines both morphology and function, physiology and pathology, as well as basic research and clinic. With this English version, this textbook is now also available for a larger audience. Anyone who has followed the publications on the subject of placenta in recent years will have noticed that the last decade alone has been marked by a large number of new findings that have led to fundamental changes in the understanding of pregnancy complications. However, these are often known to only a small community and rarely find their way into clinical research and even more rarely into clinical practice. This is where we see the great opportunity for our book. It is clear that a textbook will always be too slow to publish all these innovations directly. However, a textbook can summarize the latest insights and relate them to each other. Moreover, it can present many disciplines together in an attempt to link them together. Examples include morphology and ultrasound or function and predictive biomarkers. Our hope now is that we have succeeded in putting together a textbook that meets many demands—and from different fields and areas. We are sure that you as an

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Foreword

obstetrician and midwife, as a pathologist or basic researcher will each find important suggestions and new insights for your work. As editors, we would like to thank all the co-authors for their commitment and the publisher for his continuous support. We are pleased to have realized our dream and hope you enjoy reading it. Yours, Berthold Huppertz

Graz, Austria Ekkehard Schleußner

Jena, Germany December 2017

VII

Contents 1

Placental Development with Histological Aspects.................................................. 1 Berthold Huppertz

1.1 Introduction........................................................................................................................................... 3 1.2 Development of the Placenta.......................................................................................................... 3 1.2.1 Prelacunar Phase (Days 5 to 8 after fertilization)..............................................................................3 1.2.2 Lacunar Phase (Days 8 to 13 after fertilization).................................................................................4 1.2.3 Early Villous Phase (Days 13 to 28 p. c.)................................................................................................4 1.2.4 Villous Phase (Day 28 p. c. to end of pregnancy)..............................................................................6 1.3 Placental Villi.......................................................................................................................................... 7 1.3.1 General Histological Structure................................................................................................................7 1.3.2 Syncytiotrophoblast....................................................................................................................................8 1.3.3 Villous Cytotrophoblast (Langhans Cell).............................................................................................11 1.3.4 Villous Stroma................................................................................................................................................12 1.3.5 Placental Blood Vessels..............................................................................................................................12 1.4 Architecture of the Villous Tree....................................................................................................... 12 1.4.1 Mesenchymal Villi........................................................................................................................................13 1.4.2 Immature Intermediate Villi......................................................................................................................14 1.4.3 Stem Villi..........................................................................................................................................................15 1.4.4 Mature Intermediate Villi...........................................................................................................................15 1.4.5 Terminal Villi...................................................................................................................................................16 1.5 Extravillous Trophoblast.................................................................................................................... 16 1.5.1 Interstitial Trophoblast...............................................................................................................................18 1.5.2 Endoglandular Trophoblast......................................................................................................................20 1.5.3 Endovascular Trophoblast.........................................................................................................................21 1.5.4 Endolymphatic Trophoblast.....................................................................................................................22 1.5.5 General Considerations on Trophoblast Invasion............................................................................22 1.5.6 Maternal Perfusion of the Placenta.......................................................................................................23 1.5.7 Formation of the Chorion laeve (fetal membranes)........................................................................25 References......................................................................................................................................................25 2

Immunology of the Fetomaternal Border..................................................................... 29 Udo R. Markert, Johanna Seitz, Theresa Hofmann, Juliane Götze, and Sebastian Schamberger

2.1 Background............................................................................................................................................ 30 2.2 Immunology of the Endometrium................................................................................................. 30 2.3 Immunology of Pregnancy............................................................................................................... 31 2.3.1 Problems.........................................................................................................................................................31 2.3.2 Fetomaternal Interfaces.............................................................................................................................32 2.3.3 Pregnancy: A Th2 Phenomenon.............................................................................................................33 2.3.4 Regulatory T Cells.........................................................................................................................................34 2.3.5 γδ-T Cells..........................................................................................................................................................34 2.3.6 Natural Killer Cells........................................................................................................................................34 2.3.7 Uterine CD14+ Cells.....................................................................................................................................35 2.3.8 Trophoblastic Immunoregulatory Factors..........................................................................................36

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2.3.9 Hormones.......................................................................................................................................................37 2.3.10 Other Selected Pregnancy-promoting Mechanisms.......................................................................37 2.4 Summary and Conclusion................................................................................................................. 38 References......................................................................................................................................................39 3

Placental Morphology................................................................................................................ 43 Berthold Huppertz and Thomas Stallmach

3.1 Morphology of the Placenta............................................................................................................. 44 3.1.1 Introduction...................................................................................................................................................44 3.1.2 Villous Development...................................................................................................................................44 3.1.3 Development of Cell Columns for Trophoblast Invasion...............................................................49 3.1.4 Structures on the Villous Surface............................................................................................................53 3.2 Histopathology of the Placenta for Gynecologists................................................................... 54 3.2.1 Introduction...................................................................................................................................................54 3.2.2 First Trimester (Abortion)..........................................................................................................................54 3.2.3 Second Trimester (Hydrops Fetalis, Infection and Inflammation)..............................................57 3.2.4 Third Trimester (Circulatory and Maturation Disorders)................................................................61 3.2.5 Postpartum Period.......................................................................................................................................67 3.3 Biobanking............................................................................................................................................. 69 3.3.1 Introduction...................................................................................................................................................69 3.3.2 Variables Affecting the Composition of a Sample............................................................................70 3.3.3 Collection or Biobank?...............................................................................................................................71 Further Readings..........................................................................................................................................75 4

Placental Function—Nutrient Transport—Gas Exchange.................................. 77 Michael Gruber, Birgit Hirschmugl, Carolin Schliefsteiner, and Christian Wadsack

4.1 General Functions of the Placenta................................................................................................. 78 4.2 Nutrient Transport Across the Placenta........................................................................................ 79 4.2.1 Transport of Lipids and Fatty Acids.......................................................................................................79 4.2.2 Transport of Glucose...................................................................................................................................80 4.2.3 Transport of Proteins and Amino Acids...............................................................................................81 4.2.4 Transport of Minerals and Trace Elements..........................................................................................83 4.3 Maternofetal Gas Exchange.............................................................................................................. 85 References......................................................................................................................................................87 5

Endocrinology of the Placenta.............................................................................................. 91 Ekkehard Schleußner

5.1 Introduction........................................................................................................................................... 92 5.2 Steroid Hormones................................................................................................................................ 96 5.2.1 Progesterone.................................................................................................................................................96 5.2.2 Estrogens.........................................................................................................................................................96 5.2.3 Glucocorticoids.............................................................................................................................................97 5.3 Peptide Hormones............................................................................................................................... 98 5.3.1 Human Chorionic Gonadotropin (hCG)...............................................................................................98 5.3.2 Leptin................................................................................................................................................................99

IX Contents

5.3.3 Corticotrophin Releasing Hormone (CRH)..........................................................................................100 5.3.4 Placental Lactogen (hPL) and Placental Growth Hormone (hPGH)...........................................102 5.3.5 Summary.........................................................................................................................................................103 References......................................................................................................................................................103 6

Teratology........................................................................................................................................... 105 Herbert Juch

6.1 Introduction........................................................................................................................................... 106 6.2 Congenital Anomalies Historically................................................................................................. 106 6.3 Congenital Anomalies Today............................................................................................................ 108 6.4 Basic Risk................................................................................................................................................. 109 6.5 Medication and Pregnancy............................................................................................................... 110 6.6 Placenta and Teratology.................................................................................................................... 112 6.6.1 The Sensitivity of the Embryo to Toxic Influences Depends on the Genotype.....................112 6.6.2 The Sensitivity of the Embryo to Toxic Influences Depends on Its Stage of Development.........................................................................................................................114 6.6.3 Different Embryotoxic Influences Affect (Embryonic) Development via Relatively Few Specific Mechanisms..............................................................................................116 6.6.4 After Exposure to Teratogens, Different Developmental Courses Are Possible in Principle.............................................................................................................................116 6.6.5 The Way in Which Toxic Influences Reach the Embryo/Fetus Depends on Their Physical and Chemical Properties.........................................................................................117 6.6.6 Dose-response Relationships Apply in Teratology as Elsewhere in Pharmacology and Toxicology..............................................................................................................................................118 6.7 Conclusion.............................................................................................................................................. 120 References......................................................................................................................................................120 7

The Effects of Legal and Illegal Drugs on Placental Function.......................... 121 Justine Fitzgerald and Ekkehard Schleußner

7.1 Introduction........................................................................................................................................... 122 7.2 Smoking During Pregnancy............................................................................................................. 122 7.2.1 Tobacco Ingredients....................................................................................................................................123 7.2.2 Effects on Placental Morphology...........................................................................................................123 7.2.3 Effects on Trophoblast Cells.....................................................................................................................125 7.2.4 Oxidative Stress and Endothelial Dysfunction..................................................................................125 7.2.5 Placental Transcriptome............................................................................................................................126 7.3 Alcohol During Pregnancy................................................................................................................ 126 7.3.1 Effects on Placental Morphology...........................................................................................................127 7.3.2 Effects on Trophoblast Cells.....................................................................................................................128 7.3.3 Oxidative Stress and Endothelial Dysfunction..................................................................................128 7.4 Methamphetamines and MDMA.................................................................................................... 128 7.5 Cocaine.................................................................................................................................................... 130 7.6 Opiates..................................................................................................................................................... 130 7.6.1 Placental Transfer.........................................................................................................................................131 7.6.2 Effects on Trophoblast Cells.....................................................................................................................131 7.7 Cannabis.................................................................................................................................................. 131

X

Contents

7.7.1 Placental Transfer.........................................................................................................................................132 7.7.2 Effects on Trophoblast Cells.....................................................................................................................132 References......................................................................................................................................................133 8

Placenta-Related Hemorrhage: Pathophysiology, Diagnostics, Management..................................................................................................................................... 135 Thorsten Braun, Wolfgang Henrich, Julia Knabl, Franz Kainer Renaldo Faber, Jan Pauluschke-Fröhlich, Karl-Oliver Kagan, Harald Abele, and Lars-Christian Horn

8.1 The Placenta Accreta Spectrum (PAS)........................................................................................... 137 8.1.1 Introduction...................................................................................................................................................137 8.1.2 Epidemiology................................................................................................................................................137 8.1.3 Risk Factors.....................................................................................................................................................137 8.1.4 Definition........................................................................................................................................................138 8.1.5 Pathogenesis..................................................................................................................................................139 8.1.6 Diagnosis.........................................................................................................................................................139 8.1.7 Management and Therapy.......................................................................................................................148 8.2 Placenta Praevia................................................................................................................................... 152 8.2.1 Terminology...................................................................................................................................................152 8.2.2 Morbidity and Mortality............................................................................................................................153 8.2.3 Etiology and Risk Factors...........................................................................................................................154 8.2.4 Diagnostics and Management................................................................................................................156 8.2.5 Operational Procedure...............................................................................................................................157 8.2.6 Summary.........................................................................................................................................................159 8.3 Umbilical Cord Insertion, Variations and Vasa Praevia............................................................ 159 8.3.1 Umbilical Cord Insertion, Velamentous Cord Insertion..................................................................159 8.3.2 Vasa Praevia....................................................................................................................................................161 8.4 Premature Placental Abruption...................................................................................................... 162 8.4.1 Incidence and Risk Factors........................................................................................................................163 8.4.2 Definition........................................................................................................................................................164 8.4.3 Etiology............................................................................................................................................................164 8.4.4 Clinical Signs..................................................................................................................................................165 8.4.5 Instrumental Diagnostics..........................................................................................................................165 8.4.6 Laboratory Diagnostics..............................................................................................................................167 8.4.7 Clinical Care/Management.......................................................................................................................168 8.4.8 Conclusion......................................................................................................................................................170 8.5 Primary and Secondary Tumors of the Umbilical Cord and Placenta................................. 170 8.5.1 Tumors of the Umbilical Cord..................................................................................................................170 8.5.2 Tumors of the Placenta..............................................................................................................................171 References......................................................................................................................................................176 9

Placental Imaging.......................................................................................................................... 187 Anna-Maria Dückelmann, Hans-Joachim Mentzel, Karim D. Kalache, and Dietmar Schlembach

Sonographic Assessment of the Placenta in the Second and Third Trimester and Ultrasound/MRI Morphology of the Placenta.................................................................... 188 9.1.1 Introduction...................................................................................................................................................188 9.1.2 Localization....................................................................................................................................................190 9.1

XI Contents

9.1.3 Placenta Accreta Spectrum (PAS)...........................................................................................................193 9.1.4 Echogenicity..................................................................................................................................................197 9.1.5 Maturation of the Placenta.......................................................................................................................202 9.1.6 Size and Shape..............................................................................................................................................203 9.1.7 Placental Biometry and Volumetry........................................................................................................205 9.1.8 Summary.........................................................................................................................................................207 9.2 Doppler Sonography/Functional Diagnostics............................................................................ 208 9.2.1 Placental Vascular System.........................................................................................................................208 References......................................................................................................................................................212 10

Disorders of Early Pregnancy and Pregnancy Loss................................................. 219 Stephanie Pildner von Steinburg, Ekkehard Schleußner, Ruben Kuon, Kilian Vomstein, and Bettina Toth

10.1 Early Pregnancy and Its Disturbance............................................................................................. 220 10.1.1 Diagnosis of Early Pregnancy...................................................................................................................220 10.1.2 Pregnancy Loss.............................................................................................................................................224 10.2 Recurrent Pregnancy Loss................................................................................................................. 229 10.2.1 Introduction...................................................................................................................................................229 10.2.2 Established Risk Factors.............................................................................................................................230 10.2.3 Possible New Risk Factors.........................................................................................................................234 References......................................................................................................................................................238 11

Placental Insufficiency/Placenta-Associated Diseases......................................... 243 Berthold Huppertz, Ulrich Pecks, and Holger Stepan

11.1 Placental Disorders: Pathophysiology.......................................................................................... 244 11.1.1 Introduction...................................................................................................................................................244 11.1.2 Preeclampsia..................................................................................................................................................244 11.2 FGR: Diagnostics and Management............................................................................................... 254 11.2.1 Terminology and Definition.....................................................................................................................254 11.2.2 Epidemiology................................................................................................................................................257 11.2.3 Cause and Risk Factors...............................................................................................................................257 11.2.4 Fetal Compensation....................................................................................................................................257 11.2.5 Outcome of the Child.................................................................................................................................259 11.2.6 Diagnostics and Monitoring....................................................................................................................260 11.2.7 Therapeutic Management–Delivery Indication................................................................................263 11.3 Preeclampsia: Diagnosis and Management................................................................................ 266 11.3.1 Definition and Classification....................................................................................................................267 11.3.2 Causes and Risk Factors.............................................................................................................................267 11.3.3 Diagnosis and Early Detection................................................................................................................268 11.3.4 Biomarkers in Diagnostics and Prediction..........................................................................................269 11.3.5 Risk Assessment for Preeclampsia in the Context of First Trimester Screening and Secondary Prophylaxis......................................................................................................................270 11.3.6 Clinical Management..................................................................................................................................271 11.3.7 Possible Future Therapeutic Approaches............................................................................................272 11.3.8 Long-Term Morbidity..................................................................................................................................273 References......................................................................................................................................................274

XII

12

Contents

The Placenta in Twins.................................................................................................................. 281 Isabel Couck, Anke Diemert, Kurt Hecher, and Liesbeth Lewi

12.1 Introduction........................................................................................................................................... 282 12.2 Structural Differences Between Monochorial and Dichorial Placentas............................. 282 12.2.1 Chorionicity....................................................................................................................................................282 12.2.2 Assessment of the Placenta in Early Pregnancy................................................................................283 12.2.3 Assessment of the Placenta in Late Pregnancy.................................................................................284 12.2.4 Assessment of the Placenta after Delivery..........................................................................................285 12.3 The Placenta in Twin-to-Twin Transfusion Syndrome (TTTS)................................................ 285 12.3.1 Vascular Anastomoses in TTTS................................................................................................................285 12.3.2 Importance of the Umbilical Cord in TTTS..........................................................................................287 12.3.3 Unequal Division of the Placenta in TTTS............................................................................................287 12.3.4 Significance of Other Placental Factors for TTTS..............................................................................287 12.4 Monochorial Placenta and Discordant Growth.......................................................................... 287 12.4.1 Unequal Division of the Placenta and Discordant Growth...........................................................288 12.4.2 Anastomoses and Discordant Growth..................................................................................................289 12.4.3 Umbilical Cord and Growth Discordance............................................................................................290 12.4.4 Molecular Changes and Discordant Growth......................................................................................290 12.4.5 Other Placental Factors and Discordant Growth..............................................................................290 12.5 Dichorial Placenta and Discordant Growth................................................................................. 291 12.5.1 Umbilical Cord and Discordant Growth...............................................................................................291 12.5.2 Placental Pathology and Discordant Growth.....................................................................................291 12.6 Conclusion.............................................................................................................................................. 291 References......................................................................................................................................................292 13

Fetal Programming....................................................................................................................... 295 Evelyn Annegret Huhn, Anke Diemert, Ekkehard Schleußner, Kurt Hecher, and Petra Clara Arck

13.1 Introduction........................................................................................................................................... 296 13.2 Between Hypothesis and Epidemiology...................................................................................... 296 13.2.1 This Is How It All Began: The Barker Hypothesis and First Epidemiological Studies............296 13.2.2 Obesity, Insulin Resistance and Metabolic Syndrome....................................................................299 13.2.3 Cardiovascular Diseases.............................................................................................................................300 13.2.4 Altered Immune Response and Autoimmune Diseases................................................................301 13.2.5 Memory and Psychiatric Disorders........................................................................................................302 13.2.6 Gender-specific Programming................................................................................................................303 13.3 Underlying Mechanisms.................................................................................................................... 303 13.3.1 Direct Mother-to-Child Mediators.........................................................................................................303 13.3.2 Epigenetic Changes....................................................................................................................................307 13.4 Examples of Exogenous Stimuli of Fetal Programming.......................................................... 308 13.4.1 Maternal Malnutrition and Placental Insufficiency..........................................................................308 13.4.2 Maternal Oversupply and Gestational Diabetes...............................................................................309 13.4.3 Glucocorticoid Administration for Lung Maturation Induction..................................................309 13.5 Contradictions and Alternative Approaches.............................................................................. 310 13.6 Pregnancy as an Option for Future Health Prevention........................................................... 310 References......................................................................................................................................................311

XIII Contents

14

Fetal Cells and Cell-Free Nucleic Acids in Maternal Blood: Genetic and Immunological Aspects................................................................................ 317 Olav Lapaire, Shane Vontelin van Breda, Lenka Vokalova, Peter Celec, Irene Hösli, Simona Rossi, and Sinuhe Hahn

14.1 Introduction........................................................................................................................................... 319 14.2 Fetal Cells and Fetal DNA in Maternal Blood: A Centuries-Old Phenomenon................. 319 14.3 Cell-Free DNA: Current Status for Non-invasive Prenatal Testing........................................ 321 14.3.1 Whole Genome Sequencing....................................................................................................................321 14.3.2 Targeted Genome Sequencing...............................................................................................................322 14.3.3 SNP Approach...............................................................................................................................................324 14.3.4 Technical Principles of Currently Available Non-invasive Prenatal Tests..................................325 14.3.5 Clinical Use of NIPT, Its Limitations and Impact on Prenatal Care..............................................325 14.4 Trophoblast Cells in the Cervix – Diagnostic Use and Insight into Fetomaternal Pathologies........................................................................................................ 325 14.5 Placental Cell-Free DNA: An Activator of the Maternal Immune System and Initiator of Birth?.......................................................................................................................... 327 14.6 MicroRNA from the Placenta: New Antiviral Agents?.............................................................. 328 14.7 Circulating Fetal Cells: Their Role in Pregnancy-Associated and Postpartum Diseases.................................................................................................................. 330 14.8 Conclusions............................................................................................................................................ 331 References......................................................................................................................................................331 15

Research Aspects and In Vitro Models............................................................................. 333 Martin Gauster, Michael Gruber, Birgit Hirschmugl, Carolin Schliefsteiner, and Christian Wadsack

15.1 Trophoblast Cell Culture Models..................................................................................................... 334 15.1.1 Primary Trophoblasts..................................................................................................................................334 15.1.2 Trophoblast Cell Lines................................................................................................................................334 15.2 Placental Explant Cultures................................................................................................................ 336 15.3 Endothelial Cells................................................................................................................................... 338 15.3.1 Human Umbilical Vein Endothelial Cells (HUVECs)..........................................................................338 15.3.2 Primary Placental Endothelial Cells.......................................................................................................339 15.4 Placental Macrophages...................................................................................................................... 341 15.4.1 Decidual Macrophages..............................................................................................................................341 15.4.2 Hofbauer Cells...............................................................................................................................................341 15.5 Placenta Ex Vivo Perfusion................................................................................................................ 343 15.5.1 Methodology.................................................................................................................................................344 15.5.2 Applications in Research...........................................................................................................................346 References......................................................................................................................................................346

XIV

16

Contents

Maternal Disease Affecting the Placenta: Diabetes Mellitus............................ 349 Tanja Groten

16.1 Definition and Epidemiology........................................................................................................... 350 16.1.1 Definition........................................................................................................................................................350 16.1.2 Epidemiology................................................................................................................................................350 16.2 Effects of Hyperglycemia in Pregnancy........................................................................................ 351 16.3 Diagnosis and Therapy....................................................................................................................... 352 16.3.1 Screening and Diagnosis of Gestational Diabetes...........................................................................352 16.3.2 Therapy............................................................................................................................................................353 16.3.3 Timing of Delivery in Patients with Diabetes.....................................................................................354 16.4 Significance of the Placenta for Glucose Metabolism in Pregnancy................................... 354 16.5 Placental Changes in Patients with Diabetes............................................................................. 356 16.5.1 Placental Changes in Pre-Existing Diabetes.......................................................................................357 16.5.2 Placental Changes in Gestational Diabetes........................................................................................358 16.6 Placental Histology in Diabetes—Consequences of a Histopathological Reprocessing?........................................................................................................................................ 359 References......................................................................................................................................................361



Supplementary Information



Index................................................................................................................................................................. 365

XV

Editors and Contributors About the Editors Berthold Huppertz

has been studying the human placenta, especially the trophoblast, and its alterations in IUGR and preeclampsia for more than two decades.

Ekkehard Schleußner

is an obstetrician working on placental insufficiency and its clinical consequences. The focus of the placental laboratory of the Department of Obstetrics is placental physiology in the placental perfusion model and immunology at the maternoplacental interface.

Contributors Harald Abele, MHBA  Department of Women’s Health Tübingen, University Women’s Hospital Tübingen, Tübingen, Germany Petra Clara Arck  Center for Obstetrics, Pediatrics and Adolescent Medicine, Clinic for Obstetrics and Prenatal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany Thorsten Braun  Obstetrics Clinics, Campus Virchow Klinikum, Department of Experimental Obstetrics, Charité Universitätsmedizin Berlin, Berlin, Germany Peter Celec  Comenius University Bratislava, Faculty of Medicine, Institute of Molecular Biomedicine, Bratislava, Slovakia Isabel Couck, MD  Department of Obstetrics and Gynecology, University Hospitals Leuven, KU Leuven, Leuven, Belgium Department of Development and Regeneration, Leuven, Belgium

XVI

Editors and Contributors

Anke Diemert  Center for Obstetrics, Pediatrics and Adolescent Medicine, Clinic for Obstetrics and Prenatal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany Anna-Maria Dückelmann  Clinic for Gynecology and Obstetrics, Charité University Medical Center Berlin, Campus Virchow Clinics, Berlin, Germany Renaldo Faber  Center for Prenatal Medicine Leipzig, Leipzig, Germany Justine Fitzgerald  Center for Ambulatory Medicine I, University Clinic Jena, Eisenberg, Germany Martin Gauster  Division of Cell Biology, Histology and Embryology, Gottfried Schatz Research Center, Medical University of Graz, Graz, Austria Juliane Götze  Gutenberg Center, Mainz, Germany Tanja Groten  Clinic for Obstetrics, University Medical Center Jena, Competence Center for Diabetes and Pregnancy, Jena, Germany Michael Gruber, MSc  University Clinic for Gynecology and Obstetrics, Medical University of Graz, Graz, Austria Sinuhe Hahn, PhD  Department of Biomedicine, Laboratory of Prenatal Medicine, University Hospital Basel/University of Basel, Basel, Switzerland Kurt Hecher  Center for Obstetrics, Pediatrics and Adolescent Medicine, Clinic for Obstetrics and Prenatal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany Wolfgang Henrich  Clinic for Gynecology and Obstetrics, Charité Universitätsmedizin Berlin, Campus Virchow Klinikum, Berlin, Germany Birgit Hirschmugl, PhD  Department of Obstetrics and Gynecology, Medical University of Graz, Graz, Austria Theresa Hofmann, MD  Department of Obstetrics, Placenta Laboratory, University Hospital Jena, Jena, Germany Lars-Christian Horn  Institute of Pathology, Gyneco-, Mamma- & Perinatal Pathology Group, University Hospital Leipzig AöR, Leipzig, Germany Irene Hösli  Clinic for Obstetrics and Pregnancy Medicine, University Hospital Basel, Basel, Switzerland Evelyn Annegret Huhn  Clinic for Obstetrics and Pregnancy Medicine, University Hospital Basel, Basel, Switzerland

XVII Editors and Contributors

Herbert Juch  Division of Cell Biology, Histology and Embryology, Gottfried Schatz Research Center, Medical University of Graz, Graz, Austria Karl-Oliver Kagan, MHBA  Department of Women’s Health Tübingen, University Women’s Hospital Tübingen, Tübingen, Germany Franz Kainer  Department of Obstetrics and Prenatal Medicine, Hallerwiese Clinic, Nuremberg, Germany Karim D. Kalache, MD  Division of Maternal-Fetal Medicine, Sidra Medicine, Women’s Clinical Management Group, Doha, Qatar Julia Knabl, MD  Hallerwiese Clinic, Nuremberg, Germany Ruben Kuon, MD  Gynecological Endocrinology and Fertility Disorders, Heidelberg University Women’s Hospital, Heidelberg, Germany Olav Lapaire  University Hospital Basel, Women’s Clinic, Basel, Switzerland Liesbeth Lewi  Department of Obstetrics-Gynaecology, University Hospitals, Leuven, Belgium Department of Development and Regeneration: Pregnancy, Fetus and Neonate, University Hospitals Leuven, Leuven, Belgium Udo R. Markert  Placenta Laboratory, University Hospital Jena, Research Center Lobeda, Jena, Germany Hans-Joachim Mentzel  Institute of Diagnostic and Interventional Radiology, Section of Pediatric Radiology, University Hospital Jena, Jena, Germany Jan Pauluschke-Fröhlich  Department of Women’s Health Tübingen, University Women’s Hospital Tübingen, Tübingen, Germany Ulrich Pecks  Department of Gynecology and Obstetrics, University Hospital Schleswig-­ Holstein Campus Kiel, Kiel, Germany Simona Rossi Department of Biomedicine, Laboratory of University Hospital Basel/University of Basel, Basel, Switzerland

Prenatal

Medicine,

Sebastian Schamberger  Clinic for Anesthesia and Intensive Care Medicine, University Hospital Jena, Jena, Germany Dietmar Schlembach  Neukölln Clinic, Perinatal Center Level I, Vivantes – Netzwerk für Gesundheit GmbH, Berlin, Germany Carolin Schliefsteiner, PhD  Department of Obstetrics and Gynecology, Medical University of Graz, Graz, Austria

XVIII

Editors and Contributors

Johanna Seitz  Department of Obstetrics, Placenta Laboratory, University Hospital Jena, Jena, Germany Thomas Stallmach  Institute of Pathology Enge, Zurich, Switzerland Holger Stepan  Clinic for Obstetrics, University Hospital Leipzig, Leipzig, Germany Bettina Toth  Department of Gynecology, University Hospital for Gynecological Endocrinology and Reproductive Medicine, Innsbruck, Austria Shane Vontelin van Breda, PhD  Department of Biomedicine, Laboratory of Prenatal Medicine, University Hospital Basel/University of Basel, Basel, Switzerland Lenka Vokalova  Comenius University Bratislava, Faculty of Medicine, Institute of Molecular Biomedicine, Bratislava, Slovakia Kilian Vomstein  Department of Gynecology, University Hospital for Gynecological Endocrinology and Reproductive Medicine, Innsbruck, Austria Stephanie Pildner von Steinburg  Joint Practice of Gynecologists, Peißenberg, Germany Christian Wadsack  Department of Obstetrics and Gynecology, Medical University of Graz, Graz, Austria

XIX

Abbreviations 11β-HSD 11-beta-hydroxysteroid drogenase

dehy-

ACTH Adrenocorticotrophic hormone AEDF Absent End-Diastolic Flow AIP Abnormally Invasive Placenta APLS Antiphospholipid Syndrome cAMP  Cyclic phate

adenosine

monophos-

CBD Cannabidiol CE

Hofbauer cell

HbF

Fetal hemoglobin

hCG Human Chorionic Gonadotropin HELLP Syndrome Hemolysis, Elevated Liver enzymes, Low Platelets (syndrome) HLA Histocompatibility antigen HPA Hypothalamus-Pituitary-Adrenal (-axis)

Chronic endometritis

CHD Chorionic cavity diameter CMV Cytomegalovirus CRH  Corticotropin Releasing Hormone CRL

HBC

Crown-rump length

CTG Cardiotocography

HUAEC Human Umbilical Artery Endothelial Cell HUVEC Human Umbilical Vein Endothelial Cell

DC Dichorial

IDO Indoleamine-2,3-dioxygenase

DHEA Dehydroepiandrosterone sulfate

IFN-γ Interferon gamma

DIC Disseminated coagulopathy

IGF Insulin-like Growth Factor

intravascular

DOHaD  Developmental Origins Health and Disease DWI

of

Diffusion weighting

EFW Estimated Fetal Weight EVT Extravillous trophoblast FACS Fluorescent Activated Cell Sorting

IL Interleukin IUFD Intrauterine Death

Fetal

IUGR Intrauterine Growth Restriction IVF In Vitro Fertilization LBW Low Birth Weight

FGR Fetal Growth Restriction

LH Luteinizing Hormone

FOAD Fetal Origins of Adult Disease

MACS  Magnetic Cell Sorting

FSH Follicle Stimulating Hormone GDM Gestational diabetes GHRF  Growth Factor

Hormone

Releasing

GLUT Glucose transporter GnRH  Gonadotropin Releasing Hormone GR Glucocorticoid Receptor HbA

Adult hemoglobin

MC Monochorial MCDA Monochorial diamniotic MCMA Monochorial monoamniotic MDMA 3,4-methylenedioxymethamphetamine METH Methamphetamine

XX

Abbreviations

MPS

Massive Parallel Sequencing

MVM

Microvillous Membrane

PID Preimplantation Genetic Diagnosis

NGS

Next Generation Sequencing

PL

NICHD  National Institute of Child Health and Human Development

Placental Lactogen

PRL Prolactin REDF

Reversed End-Diastolic Flow

NIPT Non-Invasive Prenatal Testing

RSA  Recurrent tions

Spontaneous

Abor-

NK cells

Natural Killer cells

SGA

NOS

Nitric Oxide Synthetase

SLE Systemic Lupus Erythematosus

oGTT

Oral Glucose Tolerance Test

SNP Single-nucleotide polymorphisms

p.c.

post conception

SWI

Small for Gestation Age

Susceptibility weighting

PAPP-A Pregnancy-Associated Plasma Protein A

THC Tetrahydrocannabinol TNF-α

Tumor Necrosis Factor alpha

PCOS

Polycystic Ovary Syndrome

Treg

Regulatory T cell

PED

Preserved End-Diastolic flow

TSH

Thyroid-Stimulating Hormone

PGE2

Prostaglandin E2

PGF/PlGF Placental Growth Factor

TTTS  Twin-to-Twin Transfusion Syndrome

PIBF Progesterone-Induced Blocking Factor

VEGF Vascular Endothelial Growth Factor

1

Placental Development with Histological Aspects Berthold Huppertz Contents 1.1

Introduction – 3

1.2

Development of the Placenta – 3

1.2.1 1.2.2 1.2.3 1.2.4

 relacunar Phase (Days 5 to 8 after fertilization) – 3 P Lacunar Phase (Days 8 to 13 after fertilization) – 4 Early Villous Phase (Days 13 to 28 p. c.) – 4 Villous Phase (Day 28 p. c. to end of pregnancy) – 6

1.3

Placental Villi – 7

1.3.1 1.3.2 1.3.3 1.3.4 1.3.5

 eneral Histological Structure – 7 G Syncytiotrophoblast – 8 Villous Cytotrophoblast (Langhans Cell) – 11 Villous Stroma – 12 Placental Blood Vessels – 12

1.4

Architecture of the Villous Tree – 12

1.4.1 1.4.2 1.4.3 1.4.4 1.4.5

 esenchymal Villi – 13 M Immature Intermediate Villi – 14 Stem Villi – 15 Mature Intermediate Villi – 15 Terminal Villi – 16

1.5

Extravillous Trophoblast – 16

1.5.1 1.5.2 1.5.3 1.5.4

I nterstitial Trophoblast – 18 Endoglandular Trophoblast – 20 Endovascular Trophoblast – 21 Endolymphatic Trophoblast – 22

© Springer-Verlag GmbH Germany, part of Springer Nature 2023 B. Huppertz, E. Schleußner (eds.), The Placenta, https://doi.org/10.1007/978-3-662-66256-4_1

1

1.5.5 1.5.6 1.5.7

 eneral Considerations on Trophoblast Invasion – 22 G Maternal Perfusion of the Placenta – 23 Formation of the Chorion laeve (fetal membranes) – 25

References – 25

3 Placental Development with Histological Aspects

1.1

Introduction

The placenta was already recognized and revered by the early Egyptians. It was then the Greek physician Diogenes of Apollonia in the 5th century BC who was the first to describe the function of the organ in nourishing the fetus. Aristotle (384–322 BC) then followed with the description that the fetal membranes completely enclose the fetus. It was not until the Renaissance in 1559 that Matteo Realdo Colombo introduced the term “placenta”, which is traced back to the Latin root for a flat cake.

1.2

Development of the Placenta

1.2.1

 relacunar Phase (Days 5 P to 8 after fertilization)

On the 5th day after fertilization (day 5 p. c., post conception), the first cell line of the embryo, the trophoblast, has differentiated. This has led to the formation of the blastocyst. The trophoblast cells form as a spherical envelope around the embryoblast even before implantation, forming the outer envelope of the blastocyst, the trophectoderm. In the blastocyst cavity, which is enclosed by the trophectoderm, the inner cell mass of the blastocyst, the embryoblast, is found on one side in direct contact with the cells of the trophectoderm. The trophoblast cells subsequently appear exclusively in the placenta, while the cells of the embryoblast are not only responsible for the entire structure of the embryo, but also provide other cells for the placenta (villous stroma and chorion). The umbilical cord and the amnion also arise from cells of the embryoblast. The blastocyst, which adheres to the uterine epithelium at implantation (day 6-7 p. c.) and subsequently invades the connective tissue of the endometrium, consists of 107–256 cells. The majority of these cells are

1

trophoblast cells in the trophectoderm. The most common site of implantation in the uterus is found in the upper portion of the posterior wall of the uterus, near the midsagittal axis. The symmetry of the blastocyst (top-­ bottom, right-left) is determined by the position of the embryo in the blastocyst and leads, among other things, to the attachment of only those trophoblast cells to the uterine epithelium that are in direct contact with the embryo. Abnormal orientation of the blastocyst with attachment of trophoblast cells that are not in contact with the embryo may cause abnormalities in umbilical cord insertion. These abnormal orientations appear to be more common in pregnancies from in vitro fertilization (IVF) (Baergen 2011). During implantation, the trophoblast of the blastocyst proliferates to such an extent that it becomes multilayered. During this process, the outer trophoblast cells, which adhere to the uterine epithelium and are the first to penetrate this epithelium, fuse, giving rise to the first syncytiotrophoblast, which is invasive at this stage. This initially oligonucleated, later multinucleated syncytium is formed by the fusion of individual trophoblast cells, first with each other, later only by fusion of individual cells with the syncytium. The invasive syncytiotrophoblast is the only embryonic tissue in contact with maternal tissues at this stage. However, it has now been shown that the assumption is no longer true  that only through the development of the extravillous trophoblast in the 5th week of pregnancy a further cell population comes into contact with maternal tissues. It is now becoming clear that first individual extravillous trophoblast cells come into direct contact with maternal tissues very early – at the time of the prelacunar phase (Moser and Huppertz 2017). However, further studies are necessary to clarify this issue. The mononucleated trophoblast cells remaining behind the syncytiotrophoblast are now called cytotrophoblasts to distin-

4

1

B. Huppertz

guish them from the multinucleated syncytiotrophoblast. In the area of the invasion front, the invasive syncytiotrophoblast does not show a smooth surface, but is rather characterized by finger-shaped outgrowths that reach into the maternal tissues. The syncytiotrophoblast no longer has the ability to divide and relies on fusion with the underlying cytotrophoblasts. The latter proliferate continuously, while the differentiating daughter cells eventually fuse with the syncytiotrophoblast, thus enabling its growth. Hence, the syncytiotrophoblast is a true syncytium and not a cell, it has no internal cell boundaries, and does not consist of individual syncytial units. Terms such as syncytiotrophoblasts (as plural) in a placenta or syncytial cells are therefore obsolete and should be strongly avoided. 1.2.2

 acunar Phase (Days 8 to 13 L after fertilization)

During this time, the syncytiotrophoblast grows around the entire embryo so that the placenta takes on a spherical shape at the end of this phase. At day 12 p. c. the embryo is completely grown into the endometrium of the uterus and surrounded by the placenta. The uterine epithelium at the implantation site is closed again, and a thin layer of endometrial connective tissue is found between the placenta and the uterine epithelium. Around day 8 p. c., the first vacuoles appear in the syncytiotrophoblast, which merge into larger lacunae during the following days. These lacunae are separated from each other by larger structures of the syncytiotrophoblast, so-called trabeculae, which run from the embryonic side to the maternal side of the developing placenta. The fluid-­ filled lacunae gradually flow together to form a large space, which is subsequently called the intervillous space, the space between the villi. Maternal blood will later

flow into this space to ensure the supply of the placenta and fetus with nutrients and oxygen. zz Compartmentalization of the Placenta The development of lacunae leads to compartmentalization of the placenta: 55 The embryonic side of the placenta develops into the chorionic plate 55 The trabeculae develop into the villous trunks, which on the one hand connect the chorionic plate with the decidual side of the placenta and on the other hand serve as a basis for villous development 55 The developing lateral branches of the trabeculae develop into the free-­ floating villi 55 The lacunae develop into the intervillous space 55 The maternal side develops into the basal plate with parts of the uterine tissues.

1.2.3

 arly Villous Phase (Days 13 E to 28 p. c.)

Due to the proliferation of the cytotrophoblasts, a complete layer of these cells has formed under the syncytiotrophoblast. At about day 14 p. c. mesenchymal cells grow out of the embryoblast and form another layer underneath the cytotrophoblasts. At the same time, the proliferation pressure of the cytotrophoblasts pushes the cytotrophoblasts to migrate into the syncytial trabeculae until they finally reach the maternal side of the placenta where they begin to invade the uterine tissues of the placental bed as extravillous trophoblasts (trophoblasts outside the villous structures). Due to the migration of the cytotrophoblasts, the trabeculae have now developed into structures that are surrounded by the multinuclear syncytiotrophoblast and filled

1

5 Placental Development with Histological Aspects

with mononuclear cytotrophoblasts. These structures are called primary villi and are also found as branch-like structures extending from the trabeculae. The appearance of these primary villi marks the beginning of the villous phase of placentation. Shortly thereafter, the mesenchymal cells of the extraembryonic mesoderm follow the cytotrophoblasts and also invade the trabeculae and primary villi, displacing the cytotrophoblasts from the core of the trabeculae and primary villi (. Fig.  1.1). These cells penetrate almost to the maternal end of the trabeculae but do not come into contact with maternal tissues. A thick cushion of cytotrophoblasts forms between the mesenchymal cells and the maternal cells, which later is referred to as a tropho 

blastic cell column. As the cytotrophoblasts are displaced from the core areas of the trabeculae and primary villi by mesenchymal cells, secondary villi develop. Such villi are surrounded by two layers of trophoblast (syncytiotrophoblast and cytotrophoblast) and filled with a core of mesenchymal cells. On day 18–20 p. c. vasculogenesis starts in the mesenchyme of the secondary villi, which thereby develop into tertiary villi (. Fig.  1.1) (Demir et  al. 2004). Hemangioblastic stem cells within the placental mesenchyme develop into endothelial as well as blood cells. Even at this early stage, placental macrophages are found to have developed within and from the placental mesenchyme. This early development of ves 

Primary villus Syncytiotrophoblast Cytotrophoblast

Secondary villus Syncytiotrophoblast Cytotrophoblast Villous mesenchyme Trophoblastic basement membrane Tertiary villus Syncytiotrophoblast Cytotrophoblast Villous mesenchyme Placental blood vessel Trophoblastic basement membrane

..      Fig. 1.1  Schematic representation of early villous development. The cytotrophoblast pushes into the syncytial sprout of a villus by increased proliferation in this area and gives rise to a purely trophoblastic primary villus. This is followed by the connective tissue stroma, which is separated from the trophoblastic cov-

ering by a basement membrane. Thus, a secondary villus is formed. Tertiary villi then develop with the development or sprouting of blood vessels into these villi. These tertiary villi are the villi that can then be further subdivided morphologically (. Fig. 1.5)  

6

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B. Huppertz

sels and blood cells within the placenta proceeds independently of the development of vessels and blood cells in the embryo. Hematopoiesis within the placenta is found during the first trimester, then the fetal liver takes over this function. The connection of the two vascular systems of placenta and embryo via the umbilical cord does not occur until around day 35 p. c., i.e. in the 7th week of pregnancy (Benirschke et al. 2006). Due to a pronounced branching angiogenesis, almost all villi are vascularized tertiary villi by the end of the first trimester. One of the reasons for this massive angiogenesis is the intraplacental low oxygen concentration during the first trimester. The reasons for this are described and discussed in 7 Sect. 1.5.6.

On the surface of larger villi, the syncytiotrophoblast bulges outward and forms so-called syncytial sprouts. Below these syncytial sprouts, an increased proliferation of cytotrophoblasts begins, which results in filling of syncytial sprouts with cytotrophoblasts on the inside and thus becoming villous sprouts (= primary villi). Now the mesenchyme follows and  grows into the developing villi (= secondary villi). Subsequently, angiogenesis within the villous stroma leads to the ingrowth of blood vessels into these regions (= tertiary villi). Due to a corresponding growth in length combined with a permanent sprouting of new villi, the placental villous tree grows continuously until the end of pregnancy (Benirschke et al. 2006).

 

1.2.4

zz Stratification of the Placental Barrier

 illous Phase (Day 28 p. c. V to end of pregnancy)

This type of villous development results in the following stratification (from maternal to fetal) of the placental barrier, the barrier between maternal and fetal circulation (. Fig. 1.2):

The growth of the villous tree follows a uniform principle from the first villus until the end of pregnancy:

 

IVS MEr

CT

FEn

FC

ST FEr

MC

VS eBM

..      Fig. 1.2  Schematic representation of the placental barrier. The placental barrier is the barrier for the exchange of nutrients and gases between maternal and fetal blood. In this process, nutrients and gases need to transit through  the syncytiotrophoblast as well as the endothelium of the fetal capillaries. Abbre-

tBM

viations: CT Cytotrophoblast, eBM Endothelial basement membrane; FC Fetal capillary, FEn Fetal endothelium, FEr Fetal erythrocytes, IVS Intervillous space, MEr Maternal erythrocytes, MC Mesenchymal cell, ST Syncytiotrophoblast, tBM Trophoblastic basement membrane, VS Villous stroma

7 Placental Development with Histological Aspects

55 Syncytiotrophoblast: This is clearly visible in the first trimester and is characterized by an almost continuous row of nuclei. In the third trimester it thins out significantly to reduce the diffusion distance between maternal and fetal blood. At the same time, the syncytial nuclei are arranged in groups and relocated to specific areas to optimize diffusion at the sites of placental vessels. 55 Cytotrophoblast: In the first trimester there is a complete layer of cells which lie directly under the syncytiotrophoblast and are separated from the villous stroma by a basement membrane. Due to the faster growth of the mesenchymal core of the villi compared to the proliferation rate of the villous cytotrophoblast, there is a separation of the latter cells during the further course of pregnancy. In particular, villous cytotrophoblasts are rarely found at the sites where placental vessels lie closely beneath the villous trophoblast. Thus, the diffusion distance is further reduced. 55 Basement membrane of the villous trophoblast. 55 Placental mesenchyme/connective tissue of the villi: Again, in the third trimester, there is a reduction of connective tissue at the sites where vessels directly approach the trophoblastic basement membrane. This is important to reduce the diffusion distance. 55 Basement membrane of the placental endothelium: In the first trimester the placental blood vessels are not yet surrounded by a basement membrane. This develops only in the course of pregnancy. 55 Endothelial cells: The final barrier of the placental barrier is found to be the

1

endothelial cells of the placental vessels. The endothelium of the placental capillaries acts as a passive filter and limits the intercellular transfer of molecules (Eaton et al. 1993).

1.3

Placental Villi

1.3.1

General Histological Structure

The placental villi of the human placenta all share the same basic structure, although— as will be described in 7 Sect. 1.4—different peculiarities of this structure are found during pregnancy. At the same time, the placental villi are not only the site of maternofetal and fetomaternal exchange, but also the site of most of the metabolic and endocrine activities of the placenta.  

zz Histological Structure of the Placental Villi The basic structure of the placental villi follows the following scheme (from outside to inside) (. Fig. 1.3): 55 Villous syncytiotrophoblast 55 Villous cytotrophoblast (also termed Langhans cells) 55 Trophoblastic basement membrane 55 Mesenchymal stroma/connective tissue, with –– Local connective tissue cells such as mesenchymal cells, fibroblasts, myofibroblasts and smooth muscle cells –– Migration-active cells such as macrophages (also termed Hofbauer cells) –– Placental blood vessels (arteries, veins, capillaries, sinusoids)  

8

B. Huppertz

1

Villous syncytiotrophoblast Villous cytotrophoblast Trophoblastic basement membrane Villous mesenchymal stroma Fibroblast Macrophage (Hofbauer cell) Placental blood vessel

..      Fig. 1.3  Representation of the histological structures in a placental villus. Using the example of an immature intermediate villus from the first trimester, the most important histological structures of a villus are shown. At this stage, there is (1) a still double-lay-

1.3.2

Syncytiotrophoblast

The villous trophoblast, as an epithelial-like layer enclosing the villi, consists of two layers, the inner layer of the mononuclear cytotrophoblasts and the outer layer of the multinuclear syncytiotrophoblast. This syncytiotrophoblast was formed by fusion of the early trophoblast cells of the trophectoderm of the blastocyst. After implantation and throughout gestation, this syncytium is maintained by permanent fusion of the underlying cytotrophoblasts with the syncytiotrophoblast, thus introducing new cellular material into this syncytium.

In Vitro Culture and Syncytiotrophoblast Here it must be made clear that there is no fusion of villous cytotrophoblasts with each other during pregnancy, otherwise there would be multiple layers of syncytia, which have not yet been described in humans. This is different to the situation in mouse and rat, where two layers of syncytia with one layer of cytotrophoblasts have been detected. In humans, however, only fusion events between single cells and the syncytium are

ered trophoblastic coating of syncytiotrophoblast and cytotrophoblast with underlying basement membrane and (2) the villous stroma in the villous core with mesenchymal cells, fibroblasts and macrophages as well as placental blood vessels

found after implantation and during pregnancy, but not between two single villous cytotrophoblasts. This must be taken into account when performing cell cultures  with isolated cells. When villous cytotrophoblasts are isolated from a placenta and put into culture, fusion of these cells is often described as a criterion of good isolation and culture. However, if this does not occur in vivo, how can it occur in vitro? There are various explanations for this phenomenon; however, all have not yet been proven: 55 During isolation, mononuclear fragments of the syncytiotrophoblast are also isolated and taken into culture. These and other, smaller portions of the syncytiotrophoblast may serve as an initial nuclei for fusion. Once a cytotrophoblast has fused with a fragment of the syncytiotrophoblast, it may be recognized as a “syncytium” by other cytotrophoblasts. This would then result in subsequent fusion between single cells and a ‘syncytium’—similar to the in vivo situation. 55 Due to the isolation, the villous cytotrophoblasts lose contact with the underlying basement membrane of the placental

9 Placental Development with Histological Aspects

villi. In vivo, this leads to a change in the phenotype of these cells from villous to extravillous trophoblasts. It has been shown many times that multinucleated extravillous trophoblast cells occur in the placental bed. Hence, it could be speculated that the villous cytotrophoblasts change their phenotype after isolation, become extravillous trophoblasts and subsequently fuse—similar to the in vivo situation. Even though both approaches explain the behavior of villous cytotrophoblasts in culture, there is an urgent need for clarification of this behavior, since both approaches assume fundamentally different cell populations. This significantly affects the interpretation of the results of these cell cultures.

Structure and Function of the Syncytiotrophoblast The syncytiotrophoblast is a true syncytium with an apical and a basal plasma membrane. There are no lateral cell borders in this system and thus there is only one syncytiotrophoblast in a placenta. Here it must be pointed out once again that the use of the plural for the syncytiotrophoblast of a placenta or the use of the term “syncytial cells” should be urgently avoided, as this indicates above all a lack of understanding of placental structures. As the site of highest metabolic and endocrine activity in the placenta, the main role of the syncytium is maternofetal (and fetomaternal) transfer and its control. This includes but is not limited to (Benirschke et al. 2006): 55 the active transport of amino acids and electrolytes such as sodium and calcium, 55 the facilitated transport of glucose, 55 the diffusion of water and gases, but also 55 the synthesis of many hormones such as β-hCG (beta subunit of human chorionic gonadotropin) and 55 the catabolism and resynthesis of proteins and lipids.

1

Histologically, the syncytiotrophoblast is recognized as the surface boundary of each villus. Its apical surface shows a light cover with microvilli to increase the surface area for exchange. Often, the nuclei are irregularly distributed and may occur in a row or very sporadically. Functionally, the permanent fusion of the cytotrophoblasts with the overlying syncytiotrophoblast results in a constant transfer of fresh organelles, new enzyme systems and mRNA transcripts as well as whole nuclei into the syncytiotrophoblast. This helps to maintain the high metabolic activity of the syncytium, which is necessary for transfer as well as for secretory and endocrine functions. The permanent influx of new nuclei into the syncytiotrophoblast leads to the fact that the nuclei in this system are of different ages and thus show different morphologies and chromatin contents. It has been shown repeatedly that the syncytial nuclei no longer exhibit proliferative activity. The presence of rare nuclei that may still be positive for certain proliferation markers can be explained by rapid differentiation and fusion after the last cell division of a villous cytotrophoblast. The different degrees of aging of syncytial nuclei are accompanied by different degrees of transcriptional activity of these nuclei. The freshly introduced nuclei still show a certain basic activity with regard to transcription, while this activity comes to an end in the course of the retention of the nuclei in the syncytiotrophoblast (about 3–4 weeks retention of a single nucleus within the syncytiotrophoblast).

Protrusions of the Syncytiotrophoblast Histologically (and also pathologically), protrusions of different types and sizes are noticeable on the outside of the syncytiotrophoblast. The following structures can be distinguished here (. Fig. 1.4) (Kaufmann and Huppertz 2007):  

10

1

B. Huppertz

a

b

c

..      Fig. 1.4  Schematic representation of the protrusions of the syncytiotrophoblast. The “protrusions” of the syncytiotrophoblast visible in the histological sectional image can have different causes. a The development of additional villi proceeds via sprouting of primary villi from existing villi. These syncytial sprouts are found mainly in the first half of pregnancy and are true sprouts of the syncytiotrophoblast. b In the second half of pregnancy, syncytial knots are more commonly found. These structures are also outgrowths of the syncytiotrophoblast, but this time

without involvement of the underlying cytotrophoblasts. Here, apoptotic material is packed into protrusions of the apical membrane of the syncytiotrophoblast and released into the maternal circulation as the final process of apoptosis in the villous trophoblast. c The Tenney-Parker changes are misinterpretations of flat sections through the syncytiotrophoblast. Thus, these are not protrusions, but artificial formations that arise from looking onto twodimensional sectional images of three-­dimensional structures

55 Syncytial/villous sprouts: These structures—as described in 7 Sect. 1.2.4— serve the permanent growth of the villous tree of a placenta. Later, connective tissue parts of the villous stroma also push into these structures and thus lead to further branching of the villous tree. The syncytial sprouts are characterized by loosely grouped, large oval nuclei with little heterochromatin. The morphology of these syncytial nuclei suggests that the sprouts are growing structures. 55 Syncytial knots: These structures serve to dispose of aged material and thus maintain homeostasis of the syncytiotrophoblast. The nuclei are densely

packed and often show an irregular shape. They contain large amounts of  heterochromatin up to the annular chromatin of late apoptotic nuclei. Occasionally, these nuclei can also be stained with the TUNEL assay, which shows single-strand breaks in DNA as a typical feature of late apoptosis. 55 Syncytial knots are pinched off from the syncytiotrophoblast and thus released into the maternal blood flowing through the intervillous space. From there they are flushed into the maternal circulation via the uterine veins and thus enter the first capillary bed behind the placenta: the lungs. Here, these oligonuclear structures are degraded by pulmonary macro-

 

11 Placental Development with Histological Aspects

phages, leading to the fact that they are almost undetectable in peripheral blood of the mother. 55 Flat sections through the syncytiotrophoblast/Tenney-Parker changes: The structure of the placenta was, and still is, worked out mainly by histological observation of two-dimensional sections, which is why the sections of branching villi and of transitions from one villus to another are often mistakenly regarded as syncytial sprouts or knots. These are flat sections through portions of the syncytiotrophoblast that look like protrusions of the syncytium in the sectional view. There is no uniform morphology of syncytial nuclei here, as the flat sections can involve any area of the syncytiotrophoblast. Because some pathologies change the branching pattern of placental villi and thus the frequency of flat sections, these structures have often been described and named after the names of the two first describers: Tenney-Parker changes (Tenney and Parker 1940). Although not true protrusions of the syncytiotrophoblast, these structures can still be used as indicators of changes in villous structure. Quantification to classify and differentiate between pathological cases and healthy placentas is therefore quite useful. However, differentiation between ­Tenney-­Parker changes and syncytial knots in sections of a delivered placenta is extremely difficult. Often both together are wrongly described as syncytial knots.

1.3.3

Villous Cytotrophoblast (Langhans Cell)

The epithelial layer of the villous trophoblast not only consists of the multinuclear syncytiotrophoblast, but includes a layer of mononuclear cytotrophoblasts as a second layer under the syncytium. These mononu-

1

clear villous trophoblast cells have positioned themselves under the syncytiotrophoblast early in placental development and remain there until the end of pregnancy. They mainly serve the continuous supply of fresh material for the overlying syncytiotrophoblast. Thus, many proliferating cells are found in this cell layer, and mitotic figures can be observed there as well. The post-­ proliferative daughter cells within this layer  start their differentiation, thereby significantly increasing their transcription and translation rates and thus store large amounts of mRNA and proteins, which are then transferred into the syncytiotrophoblast during the subsequent syncytial fusion event. Morphologically, villous cytotrophoblasts are characterized by being roundish in section, sometimes polygonal. Their nuclei are large and weakly stained, with little heterochromatin. The importance of maintaining the syncytiotrophoblast for the function of the villous cytotrophoblasts becomes apparent when the syncytium is damaged. Breaks in the syncytium are resealed by fibrin deposits initiated by maternal blood (fibrin-type fibrinoid, perivillous fibrin deposition). If such an area is so large that underlying villous cytotrophoblasts lose contact with the syncytiotrophoblast, these single cells start their differentiation towards the extravillous trophoblast phenotype. Thus, they are no longer available as proliferating progenitor cells for the syncytiotrophoblast. While in the first trimester there is still an almost complete layer of villous cytotrophoblasts under the syncytiotrophoblast, this layer thins out in the course of pregnancy, so that in the third trimester only individual cells are found under the syncytium. This is mainly due to an enormous expansion of the volume of the villous stroma, rather than a decrease in the proliferation activity of the cytotrophoblast. This can be seen from the fact that the ratio of nuclei in cytotrophoblasts to nuclei in the

12

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B. Huppertz

syncytiotrophoblast is about 1:9, regardless of whether this value is determined in the period 13–16 weeks of gestation or 37–41 weeks of gestation (Mayhew et al. 1999). 1.3.4

Villous Stroma

The connective tissue stroma of the placental villi develops from the  extraembryonic mesoderm of the early villous phase (7 Sect. 1.2.3). The initially predominant fixed connective tissue cells, the undifferentiated mesenchymal cells, differentiate into fibroblasts, which are the most widespread cells in the stroma by the end of pregnancy. At the end of pregnancy, the mesenchymal cells appear only in small areas, especially directly under the basement membrane of the villous trophoblast in stem villi. Around larger vessels, the fibroblasts further differentiate into myofibroblasts, which become smooth muscle cells in the media of the vessels. The fixed connective tissue cells of the placental stroma produce many connective tissue fibers, including type I and III collagens and fibronectin. These fibers serve to increase the mechanical stability of the villi. This is particularly necessary against the background of the permanent movement through the two blood flows of mother and child in the placenta. Macrophages, also called Hofbauer cells in the placenta, make up the majority of free connective tissue cells. These macrophages either originated directly from mesenchymal progenitor cells in the placenta or were recruited from the fetal circulation. To date, no marker is known that can distinguish these two subtypes of Hofbauer cells. Hofbauer cells secrete a large number of growth factors and are also involved in the differentiation of the villous trophoblast and in vasculo- and angiogenesis in the placenta (Demir et al. 2004). In addition to Hofbauer cells, a few plasma and mast cells are also found in the villous stroma.  

1.3.5

Placental Blood Vessels

Embedded in the villous stroma are the placental vessels. Starting from the two umbilical arteries, the fetal blood flows through the chorionic plate arteries into the arteries of the villi. There the arterial vessels branch further until, starting from small arterioles, the blood flows through the capillary bed of the placenta. This capillary bed with a surface of more than 10 m2 serves the exchange between mother and child. At the time of delivery, the capillary bed is mainly found in specific types of villi (Terminal villi; 7 Sect. 1.4.5). In such villi, the blood vessels are mostly found in direct vicinity of the villous trophoblast. The return flow to the fetus then takes place via the venous leg, which runs via the villous veins into the chorionic plate veins and from there into the one umbilical vein. The contractile cells around the large placental vessels are clinically important. Reduced fetoplacental perfusion, which can be visualized by Doppler ultrasound of the umbilical arteries, is closely associated with reduced fetal growth (FGR, “fetal growth restriction”, or IUGR, “intrauterine growth restriction”), which can lead to intrauterine fetal death (Krebs et al. 1996). Because placental vessels have no autonomic innervation (as the entire placenta has no innervation), regulation of placental blood flow occurs via local and systemic factors that control blood flow along with the anatomical arrangement and fetal cardiac output.  

1.4

Architecture of the Villous Tree

Even if the presentation of the architecture of the placental villous tree seems like an academic exercise, it has been shown that this architecture has clinical relevance. The significant changes in villous structure and types in conjunction with changes in the pla-

13 Placental Development with Histological Aspects

cental vasculature are clearly seen in FGR and are associated with placental insufficiency. During pregnancy, a total of five types of villi have been defined based on circumference/diameter, stromal characteristics, and vascular structures (Benirschke et  al. 2006; Baergen 2011) (. Fig. 1.5). Over the course of pregnancy, villous types of different appearance and functions develop. Thus, each stage of pregnancy can be assigned to a specific set of villous types and functions (. Fig. 1.6).  

 

1.4.1

Mesenchymal Villi

Mesenchymal villi (. Fig. 1.5a) are the first villi to develop as tertiary villi and thus have all three major components of a villus: villous trophoblast, villous stroma, and placental blood vessels. This occurs at about day 20 p. c. Thus, mesenchymal villi are the precursors of all other types of villi. In the first and second trimesters, mesenchymal villi differentiate into immature intermediate villi (7 Sect. 1.4.2), whereas in the third trimester they develop predominantly into  

 

a

c

1

b

e

d

..      Fig. 1.5  Schematic representation of the different types of villi. The five different types of villi of the human placenta can be distinguished on the basis of morphological criteria. a Mesenchymal villus with many mesenchymal cells, small vessels and a bilayered trophoblast. b Immature intermediate villus with a loose stroma, small vessels and stromal channels.

Again, the villous trophoblast is bilayered. c Large stem villus with two centrally located vessels (artery and vein), dense fibrous stroma, and only few sporadic cytotrophoblasts. d Mature intermediate villus with loose connective tissue, small vessels and few cytotrophoblasts. e Terminal villus with sinusoids and capillaries lying closely under a thin syncytiotrophoblast

14

B. Huppertz

1

Placental development Implantation

Birth

Last period

Maternal blood flow through the placenta

3rd week of gestation

12th week of gestation

40th week of gestation

Mesenchymal villi (5-23 (40) weeks gestation)

Villous growth

Immature intermediate villi (8-20 (40) weeks gestation)

Villous Stabilization

Stem villi

(18/20 to 40 weeks gestation)

Mature intermediate villi (24 to 40 weeks gestation)

Fetal nutrition

Terminal villi

(27 to 40 weeks gestation)

(from the 34th week the dominant villous type)

..      Fig. 1.6  Representation of the development of villous types during pregnancy. Over the course of pregnancy, the change in villous types becomes obvious. At the beginning, mesenchymal villi and immature intermediate villi are necessary for the growth of the villous trees, but around mid-­gestation, the villous

structures are increasingly stabilized by the developing stem villi. With the second half of pregnancy, mature intermediate villi and finally terminal villi develop for the adequate nutrition of the fetus until birth

mature intermediate villi (7 Sect. 1.4.4). While they are the predominant type of villi at the beginning of pregnancy, mesenchymal villi in a term placenta are only found in very small areas in the centers of villous trees and account for less than 1% of the volume of all villi. Mesenchymal villi are about 40–80 μm in diameter, have a distinct bilayered villous trophoblast, and have a stromal core with few extracellular fibers such as collagen type I, few fibroblasts and Hofbauer cells, and first small blood vessels. However, there are many mesenchymal cells in the stroma. Due to the high proliferation rate of mesenchymal cells and cytotrophoblasts in this type of villi, mesenchymal villi are the villi with the highest proliferation rate.

1.4.2

 

Immature Intermediate Villi

Immature intermediate villi (. Fig.  1.5b) are the first type of villi differentiating from mesenchymal villi, starting around the 8th week of gestation. Through massive growth and further differentiation, this type of villus becomes the predominant type of villus between weeks 14 and 20 of gestation. In a term placenta, however, this type of villus, similar to the mesenchymal villi, almost completely disappears and only accounts for about 5% of the volume of all villi. Like mesenchymal villi, immature intermediate villi have a distinct bilayered villous trophoblast. However, they are significantly larger with a diameter between 100 and 300 μm. Their stromal core shows loose reticular  

1

15 Placental Development with Histological Aspects

connective tissue with few matrix components, small blood vessels and a large number of Hofbauer cells. These cells stand out in immature intermediate villi because these villi have a distinctive feature not found in any other type of villus: stromal channels. These are fluid-filled cavities that run along the longitudinal axis of the villi. They are surrounded by fibroblasts but have no connection to the vasculature and also show no demarcation by endothelial cells. Hofbauer cells are often found in these channels, hence easily recognizable in this type of villus, although macrophages are present in all types of villi. The immature intermediate villi are a type of villus that is only found temporarily, as they develop from mesenchymal villi and subsequently differentiate into stem villi (7 Sect. 1.4.3). This explains the marked decrease in the number of this type of villus at the beginning of the second half of pregnancy. Immature intermediate villi are often mistakenly called edematous villi because the presence of stromal channels is misinterpreted. The retention of this type of villus in a mature placenta is not due to edematous changes in the placenta, rather it is a clear indication of a lack of villous maturation and is associated with fetal growth restriction.  

1.4.3

Stem Villi

again stem villi (second generation). Also the anchoring villi that connect the placenta to the basal plate are stem villi. Even if the caliber of this type of villus shows clear differences, the morphological characteristics are relatively clear. The villous trophoblast is no longer very pronounced in stem villi and is often replaced by fibrin deposits. The stroma is filled with many fibers of the extracellular matrix to account for the mechanical stability of these villi. The number of fibroblasts and Hofbauer cells is relatively low, and a few mast cells are also found. On the other hand, the very large villi are characterized by the presence of two central vessels (one artery, one vein). Around the media of these vessels contractile cells (myofibroblasts) are found, which represent a placenta-specific perivascular system for the local regulation of the vessel width and hence the blood flow through the villous trees. In addition, there are small vessels at the periphery of stem villi. The function of the stem villi is the mechanical stabilization of the villous trees of a placenta, and here they function similarly to the trunk of a tree. Their function is not the exchange of nutrients and gases but rather to transport nutrient and oxygen-rich blood from the villous trees to the fetus (large vein) and to transport nutrient and oxygen-depleted blood from the fetus to the villous trees (large artery) to the exchange areas of the terminal villi (7 Sect. 1.4.5).  

Stem villi (. Fig.  1.5c) are formed by further differentiation of the immature intermediate villi and appear from the 18th/20th week of pregnancy. They are the largest villi in a placenta and range in diameter from 100 μm to several millimeters and can be seen with the naked eye. Due to their size, they account for about 20-25% of the volume of all villi in a term placenta. They extend as large trunks from the chorionic plate into the intervillous space. Many of the first branches of these large trunks are  

1.4.4

Mature Intermediate Villi

At the beginning of the third trimester (approximately from the 24th week of gestation), the mesenchymal villi no longer differentiate into immature intermediate ­ villi, but into a new type of villus, mature intermediate villi (. Fig. 1.5d) with a diameter of 80–120 μm. While the differentiation into immature intermediate villi was crucial  

16

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for the general growth of the villous trees, the differentiation into mature intermediate villi is mandatory for the increased development of the exchange surface of the placenta. The latter is required to meet the significantly increased needs of the fetus in terms of nutrient and oxygen uptake. The mature intermediate villi make up about 25% of the volume of all villi in a term placenta. Mature intermediate villi are long and narrow, possessing a loose connective tissue stroma and many, often marginal, small vessels, predominantly capillaries, but also small terminal arterioles and venules. 1.4.5

Terminal Villi

Terminal villi (. Fig. 1.5e) are the last type of villi to differentiate during pregnancy. They arise from increased growth in length of the capillaries in mature intermediate villi, which bulge outwards as a result of this growth, thus generating new villi. Terminal villi are therefore not formed by the differentiation of precursor villi like intermediate and stem villi, but from the 27th week of pregnancy by protrusions starting from mature intermediate villi. The function of terminal villi is to facilitate exchange between mother and fetus across the placental barrier. Therefore, this type of villi accounts for about 40% of the villous volume in a term placenta. Since they are terminal protrusions, their length can be given as up to 200 μm, with a width of 50–100 μm. The trophoblastic coating of terminal villi varies in thickness. Where capillaries lie directly beneath the villous trophoblast, the thickness of the syncytiotrophoblast is reduced to a few micrometers and nuclei are absent, while cytotrophoblasts are very rarely found in these areas. Here the basement membrane of the capillaries comes into direct contact with the trophoblast basement membrane. Thus, at these sites the placental barrier has its least thickness with (1) the syncytiotrophoblast, which is mini 

mally thick, (2) the fused basement membranes of trophoblast and capillary, and (3) the endothelium. This unit is also referred to as the “vasculosyncytial membrane” in the literature. Here the diffusion distance is reduced to 0.5–2 μm. In areas without direct connection to underlying vessels, the syncytiotrophoblast is much thicker, nuclei often lie together in groups, and cytotrophoblasts are also found here. In terminal villi, the capillaries in the stroma dilate, becoming sinusoids to increase the exchange area and slow blood flow in these areas. Thus, the stroma of terminal villi is characterized by capillaries and sinusoids, which may account for more than 30% of the stromal sectional area of a terminal villus. The number of terminal villi adapts to the microenvironment in the placenta. In mothers with chronic anemia or in mothers living at high altitude, less oxygen reaches the placenta. The placenta in the second and third trimester responds by increasing its exchange surface area. Increased proliferation of endothelial cells in the mature intermediate villi leads to increased formation of terminal villi. On the other hand, the opposite picture is seen in growth-restricted fetuses with insufficient circulation through the placenta (AEDF, “absent end-diastolic flow”, or REDF, “reversed end-diastolic flow”, in the umbilical arteries). In these cases, too little oxygen is transported from the placenta to the fetus. This results in an increase in oxygen concentration within the placental intervillous space compared to the normal weight fetus. Again, the placenta reacts, this time with a reduction in surface area and regression of terminal villi. 1.5

Extravillous Trophoblast

During the early villous phase (about day 15 p. c.), mononuclear trophoblast cells come into direct contact with maternal tissues for the first time (7 Sect. 1.2.3). Because of  

17 Placental Development with Histological Aspects

1

their location outside the villous structures, also determines the depth of invasion, since they are called “extravillous trophoblasts”. it depends directly on the lifespan of these All trophoblasts that do not function as cells and cannot be transferred by proliferaepithelial-­like cells of the villi are grouped tion to daughter cells, which could then under the term extravillous trophoblast. expand the depth of invasion. This includes trophoblasts in the chorionic Invasion of extravillous trophoblasts is plate, in the fetal membranes, in the basal not limited to early pregnancy but is a conplate and placental bed, and in villous areas tinuous process until the end of pregnancy. where the trophoblastic covering is no lon- The area of the uterine wall that lies beneath ger present. All these extravillous tropho- the placenta and is invaded by extravillous blasts have in common that they express trophoblasts is called the placental bed. The histocompatibility antigen G (HLA-G) and portion of the decidua that is invaded is can thus be distinguished from the villous named decidua basalis. The portion of the trophoblast (McMaster et al. 1995). HLA-G decidua basalis that is delivered with the is mainly used in (immuno)histological placenta at birth is called the basal plate. examinations as an unambiguous marker for the extravillous trophoblast. zz Populations of the Extravillous After penetrating the syncytiotrophoTrophoblast blast in the early villous phase, the extravilTrophoblast invasion serves multiple purlous trophoblasts form multilayered poses and is implemented by distinct popcolumns of cells at these sites, the so-called ulations of extravillous trophoblasts trophoblast cell columns. Within these cell (. Fig. 1.7): columns, there is a gradient starting from 55 Interstitial trophoblast: origin of all the villous trophoblast on the basement other populations of the extravillous membrane of the anchoring villus to the distrophoblast in the placental bed, starttally located invasive extravillous trophoing from the trophoblast cell columns, blasts. The cells adhering to the basement this is the population found in the conmembrane show proliferative activity, which nective tissue of the decidua basalis and is lost in the next layers. The post-­ in the myometrium; function: adhesion proliferative daughter cells are pushed disof the placenta to the uterine wall. tally by proliferative pressure and change 55 Endoglandular trophoblast: erosion their phenotype. and opening of the uterine glands to The cells start to express HLA-G and allow histiotrophic nutrition of the also change their expression pattern of embryo during the first trimester. matrix proteins, matrix-degrading proteases 55 Endovascular trophoblast: So far, and integrins to start their invasion into the only the extravillous trophoblast maternal tissues of the uterus. The expresinvading the spiral arteries was called sion pattern and the invasive behavior of endovascular trophoblast. Today, it extravillous trophoblasts are very similar to appears that also the uterine veins are those of invasive tumor cells. However, they invaded. Therefore, the nomenclature differ from tumor cells in one crucial aspect: needs to be updated here. Extravillous trophoblasts that exhibit inva–– Endoarterial trophoblast: erosive behavior have lost their proliferative sion, opening and remodeling of activity. Should they be flushed into the the uterine spiral arteries to promaternal bloodstream, metastasis cannot vide hemotrophic nutrition of the occur because these cells no longer exhibit the ability to divide. At the same time, this  

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fetus during the second and third trimesters. –– Endovenous trophoblast: erosion and opening of the uterine veins to ensure the return of maternal blood from the intervillous space of the placenta back into the maternal system. 55 Endolymphatic trophoblast: Also the uterine lymphatic vessels are eroded by invading extravillous trophoblasts. The purpose of this erosion is not yet clear.

1.5.1

Interstitial Trophoblast

The cells that initially invade the uterine connective tissue are called interstitial extravillous trophoblasts and are thus precursors of all other types of extravillous trophoblast in the placental bed. These cells invade down into the inner third of the myometrium and thus do not reach the outer wall of the uterus in a normal pregnancy. On their way through the uterine interstitium, the interstitial trophoblasts secrete extracellular matrix proteins, which in their entirety are named matrix-type fibrinoid

Anchoring villus Trophoblastic cell column Villous trophoblast Extravillous trophoblast Interstitial trophoblast -> Connective tissue of the placenta (Decidua basalis and inner third of the myometrium) Endoglandular trophoblast -> Uterine glands Endovenous trophoblast -> Uterine veins Endoarterial trophoblast -> Uterine spiral arteries

Endovascular trophoblast -> Uterine vessels

Endolymphatic trophoblast -> Uterine lymphatic vessels

..      Fig. 1.7  Representation of the differentiation of the extravillous trophoblast. Starting from the cell columns of the anchoring villi, extravillous trophoblasts differentiate and migrate into maternal uterine tissues. During this process, they reach the luminal structures of the uterine wall. It is now apparent that e­ xtravillous

trophoblasts invade all luminal structures in the uterus, although an explanation of why this occurs is still pending, at least in the case of lymphatic vessels. At the same time, the nomenclature had to be changed to represent the invasion of all luminal structures in a meaningful way

19 Placental Development with Histological Aspects

(Kaufmann et  al. 1996) and serve as glue for anchoring the placenta to the uterine wall. The population of the interstitial trophoblast is morphologically and functionally very heterogeneous. Different phenotypes can be distinguished with respect to invasion behavior, contact to maternal cells, and secretion of and attachment to extracellular matrix components. So far, the molecular basis for these different phenotypes is missing; however, there are corresponding explanations for the different trophoblast phenotypes in the mouse (Simmons et al. 2007). A better understanding of the molecular basis of trophoblast phenotypes in humans would help to understand the deviations from them in specific diseases during pregnancy. These include premature placental abruption, fetal growth restriction (FGR) and early preeclampsia. Typically, three phenotypes can be distinguished morphologically and functionally in the population of interstitial trophoblasts: 1. Large polygonal cells Morphologically, these cells are large, polygonal cells with a single nucleus that is large, irregularly shaped, and clearly stainable. No other phenotype of all trophoblasts shows stronger immunohistochemical staining for cytokeratin 7. At the same time, these cells are always negative for proliferation markers such as Ki-67. This phenotype was previously described as X cells and is the cell type that predominantly secretes matrix-type fibrinoid (Kaufmann et  al. 1996). The matrix contains components of basement membranes such as laminins and type IV collagen, but also fibronectin and vitronectin and an amorphous ground substance containing heparan sulfate (Huppertz et  al. 1996). These large cells surround themselves with this

1

matrix and anchor to this matrix with appropriate integrins (alpha5/beta1, alpha1/beta1 and alpha-v/beta3/5 integrins). Thus, they are rarely in direct contact with maternal cells and can be detected in the uterus even decades after the last pregnancy. The above characteristics of these cells do not indicate a strong invasiveness of these cells. Rather, this phenotype is thought to perform the task of attaching the placenta to the uterine wall. This also explains the strong expression and secretion of matrix-type fibrinoid by these cells. In early pregnancy (week 9–12), this phenotype accounts for about 45% of all interstitial trophoblasts. This proportion increases during pregnancy to 69% (week 16–24) and peaks at term at 89% (week 31–39) (Benirschke et  al. 2006). These cells are relatively evenly distributed over the basal plate and the depth of invasion, reaching the inner third of the myometrium. 2. Small spindle-shaped cells There are only few descriptions of this phenotype so far. One of the reasons for this could be that these cells are often overlooked when looking at tissue sections due to their morphology, which is very small and spindle-­shaped. This phenotype is only moderately immunoreactive for cytokeratin 7 and is—like all extravillous trophoblasts—negative for proliferation markers such as Ki-67. Structurally, this phenotype is characterized by small, oval nuclei and its elongated, almost filamentous cell body, which is oriented in the direction of invasion and surrounded by only little matrix. These small cells secrete little extracellular matrix, which consists mainly of cellular and oncofetal fibronectin. At the same time, these cells express only interstitial integrins (such as alpha5/beta1

20

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B. Huppertz

and alpha-v integrins). This combination of and interaction between interstitial integrins and oncofetal fibronectins is an essential mechanism for trophoblast invasion (Huppertz et  al. 1996). This phenotype is thought to be highly invasive. Contrary to the large polygonal phenotype, the number of the small phenotype is reduced during pregnancy. While its proportion is about 55% during early pregnancy, it is only 31% at mid gestation and further reduced to 11% of the interstitial trophoblasts at term (Benirschke et al. 2006). This could be an indication of the decreasing invasive activity of the extravillous trophoblast and the decreasing proliferation activity of the trophoblast cell columns. These small cells can be visualized over the entire invasion area and thus show the same distribution pattern as the large polygonal cells. 3. Multinuclear giant cells The difference to the other phenotypes of the interstitial trophoblast is that these cells have more than one and up to ten nuclei. These nuclei are irregularly shaped and have different sizes. Thus, these cells have a much larger volume than the other phenotypes, with cell diameters between 50 and 100 μm (Benirschke et  al. 2006). The divisionincompetent multinuclear giant cells are either mostly immunonegative for cytokeratin 7 or show only small areas of immunoreactivity. Thus, they can be easily overlooked in a superficial view of trophoblast invasion. Although syncytial fusion of extravillous trophoblasts has never been observed, it is generally accepted that these giant cells result from fusion of mononuclear single cells of the interstitial trophoblast. This phenotype is found predominantly in the depth of the placental bed, at the boundary between the decidua and the myome-

trium. Its share in the total number of interstitial extravillous trophoblasts is very small. 1.5.2

Endoglandular Trophoblast

In recent years, the picture of trophoblast invasion has changed significantly. So far, only two types of extravillous trophoblast have been described, the interstitial trophoblast and the trophoblast surrounding  and invading spiral arteries. Today, the picture has been supplemented by several new populations of the extravillous trophoblast (Moser et al. 2010, 2015). These include the endoglandular trophoblast, which, differentiating from the interstitial trophoblast, reaches and erodes the uterine glands early in gestation. Secretion products from the uterine glands in the intervillous space of the placenta in the first trimester of pregnancy were described earlier, but without knowing the mechanisms by which the secretion products entered this space (Burton et  al. 2002, 2007). Only the demonstration that extravillous trophoblasts not only migrate past the uterine glands but specifically invade them, clarified how histiotrophic nutrition of the embryo can be achieved before the start of maternal blood flow into the placenta (Moser et al. 2010). Infiltration into the glands and replacement of the glandular epithelium results in opening of uterine glands by endoglandular trophoblasts, creating the pathway to connect these luminal structures of the uterus to the ­intervillous space of the placenta (Moser et al. 2010). Recent studies expand this picture: close examination of archival material from the earliest placental stages shows that the endoglandular trophoblast is the first population of the extravillous trophoblast to erode uterine  luminal structures during pregnancy (Moser and Huppertz 2017).

21 Placental Development with Histological Aspects

1.5.3

Endovascular Trophoblast

For decades it was assumed that only the spiral arteries were opened and remodeled by extravillous trophoblasts. Invasion into uterine veins was always excluded. This did not take into account that maternal blood flowing into the placenta via the opened spiral arteries needs to be transported back into the maternal bloodstream via maternal veins in the uterus. The discovery of trophoblast invasion into uterine veins was therefore not unexpected. However, this fact also implies that the term “endovascular trophoblast” no longer refers only to cells that invade arteries (Kaufmann et  al. 2003). Meanwhile, this term must be broadened to serve for all extravillous trophoblasts that invade vascular structures of the mother (i.e., arteries and veins). Thus, new terms need to define specifically invading trophoblast populations. All extravillous trophoblasts that invade vascular structures are now grouped together as endovascular trophoblasts. The cells that invade arteries are called endoarterial trophoblasts, while the cells that invade veins are named endovenous trophoblasts (Moser and Huppertz 2017).

Endoarterial Trophoblast Starting from the interstitial trophoblast, a specific population of trophoblasts reaches the uterine spiral arteries, penetrates their media and also their endothelium and thus enters the lumen of these vessels. Contrary to some notions, this endoarterial trophoblast (formerly endovascular trophoblast) must penetrate the vessel wall from the outside to reach the interior of the vessels (intravasasion). The notion that these trophoblasts start to erode the walls of the vessels from within must be abandoned, since no cells, and thus no basis, exist for such extravasasion. It is quite conceivable that endoarterial cells, once invaded from the interstitium into the vessel, migrate  along

1

the luminal side of the endothelium of the vessel wall and subsequently re-invade the vessel wall (Kaufmann et al. 2003). The main task of the endoarterial trophoblast is not only to open the spiral arteries towards the intervillous space of the placenta, but above all the conversion of the arteries into dilated tubes, which are no longer subject to maternal vascular control. This, however, refers only to the terminal parts of these vessels lying furthest towards the placenta in the region of the decidua. Thus, invasion into arteries is the only trophoblast invasion in which, in addition to the opening of a luminal structure, the wall of these structures is also remodeled. In the case of invasion into glands, veins and lymphatic vessels, these structures are only opened, but the walls are not further remodeled. The transformation of the spiral arteries can be divided into three stages: 1. Maternal factors induce initial changes in the uterine spiral arteries very early in pregnancy, long before the first extravillous trophoblast has reached these vessels. The vessels begin to dilate due to decreasing vascular smooth muscle cell organization and altered endothelial cell morphology. It appears to be maternal uterine immune cells that are responsible for these changes. They accumulate in the decidua around these vessels and play an active role in the early stages of remodeling of these vessels. In this process, they secrete growth factors and matrix metalloproteinases that degrade the extracellular matrix of the vessel walls. 2. Once the vessel wall is disorganized and the vessel is slightly dilated, endoarterial trophoblasts from the connective tissue of the decidua invade and penetrate the vessel wall. The cells that have migrated into the vessel lumen form a plug that closes these vessels, preventing maternal blood from flowing into the placenta (Weiss et al. 2016).

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3. Endoarterial trophoblasts subsequently also invade the vessel wall in the deeper areas of the vessels, and some of the cells that have already invaded will probably also migrate down the endothelium and may invade into the vessel wall again. Thus, the vessel walls are further invaded from the outside and inside and thus disorganized. This leads to further dilatation of the vessels, which in turn results in a vessel diameter many times larger than that of the original vessel. Reduced smooth muscle cell activity and loss of elastic fibers further contribute to the increase in vessel diameter.

Endovenous Trophoblast Invasion of extravillous trophoblasts into uterine veins has only been described for the first time in 2017 (Moser et  al. 2017). As outlined in 7 Sect. 1.5.3, there must be a mechanism that also connects the uterine veins to the intervillous space to allow maternal blood flow from the placental intervillous space back into the mother’s bloodstream. Thus, the new concept of venous invasion does not challenge the already existing concept of endoarterial (formerly endovascular) trophoblast invasion, but extends it in a useful way. While invasion into uterine spiral arteries involves not only opening and connection to the intervillous space, but also remodeling of the vessel wall and plug formation in the lumen of these vessels, only opening and connection to the intervillous space seem to be crucial for invasion of the uterine veins—as is the case for the glands. Endovenous trophoblasts are found in the immediate vicinity of the veins, replacing the endothelial cells of the veins, and also in the lumen of these vessels. However, so far there is no evidence of remodeling of the vessel walls and no evidence of occlusion of the lumen. Neither would make sense functionally, since reflux into the maternal system needs to occur very early in pregnancy.  

1.5.4

Endolymphatic Trophoblast

In the meantime, there are first indications that the uterine lymphatic vessels are also eroded. Two publications from 2017 show first data on this (He et al. 2017; Windsperger et al. 2017). Why invasion into these luminal structures of the uterus occurs is still unclear. It is possible that the extravillous trophoblast is more invasive than previously thought and invades all luminal structures that lie within its expansion range.

1.5.5

General Considerations on Trophoblast Invasion

The invasive potential of the extravillous trophoblast is optimally regulated. Thus, only few malignant transformations of the extravillous trophoblast have been described so far. Chorio carcinomas arise from villous but not from extravillous trophoblasts. For further information, see Sect. 3.2 “Histopathology of the placenta”. z Excessive Invasion

Alterations on the maternal side, such as uterine scars, can lead to dramatic changes in invasion. In such cases, extravillous trophoblasts may invade deeper than usual (Jauniaux et al. 2016): 55 Placenta accreta: Here the decidua is partially or completely absent, so that the placenta is firmly attached to the uterus. 55 Placenta increta: Here the trophoblast invasion may extend far into the myometrium. 55 Placenta percreta: Here the extravillous trophoblasts invade through the entire wall of the uterus and may even infiltrate the bladder wall (in the case of an anteriorly located placenta). In all three situations, placental delivery after birth is often difficult or impossible

23 Placental Development with Histological Aspects

and is associated with massive maternal bleeding. Reasons for the increasing number of placenta accreta/increta/percreta cases have now been identified. With an increase in cesarean sections in previous births (Silver et al. 2006) as well as after successful endoscopic surgery for Asherman syndrome (Fernandez et  al. 2006), the risk of deeper trophoblast invasion also increases. This may indicate that the myometrium arrests trophoblast invasion. In areas where the musculature is absent, e.g. due to scar tissue, this blockade is absent, resulting in deeper trophoblast invasion (Sect. 8.1). z Insufficient Invasion

On the other hand, insufficient invasion and thus anchorage of the placenta to the uterine wall can lead to premature detachment of the placenta, either before birth or during labor. An almost complete absence of the endoarterial trophoblast characterizes the placental bed of miscarriages that occur late in the first trimester and have chromosomally normal fetuses (Ball et  al. 2006a). Interestingly, no irregularities in trophoblast invasion were found in very early miscarriages (Ball et al. 2006b). If the endoarterial trophoblast fails to occlude all arteries connected to the placenta during the first trimester, local influx of oxygen-rich maternal blood into the intervillous space may occur too early. This has dramatic consequences for the corresponding area of the placenta, such as a marked reduction in trophoblast proliferation and decreased villous angiogenesis. This leads to disruption of placental development resulting in miscarriage in severe cases. Mild cases may result in severe fetal growth restriction (FGR) of the fetus with or without preeclampsia (Kadyrov et  al. 2003; Jauniaux et al. 2003b). The premature influx of maternal blood into the placenta can be clinically demonstrated by transvaginal ultrasound (Jauniaux et al. 2003b).

1

These clinically relevant deviations from the norm highlight the importance of understanding extravillous trophoblast invasion in detail. 1.5.6

Maternal Perfusion of the Placenta

The perfusion of the placenta with maternal blood differs fundamentally between the first trimester and the two later thirds of pregnancy. In the first trimester of pregnancy, maternal perfusion takes place with an ultrafiltrate of blood without blood cells (blood plasma), but with secretion products of the uterine glands (histiotrophic nutrition of the embryo). During the other two thirds of pregnancy, the placenta is perfused with maternal blood (hemotrophic nutrition of the fetus).

In the First Trimester Already at the time of the early villous phase, the first superficial capillaries of the decidua are eroded by the invasion of the trophoblast, which then flush a few blood cells into the developing intervillous space. However, these few blood cells do not set up a maternal circulation through the placenta at this time of pregnancy. Later during the first trimester, the spiral arteries are opened towards the placenta by the endoarterial trophoblast but are directly occluded before an influx of maternal blood into the placenta can occur (Burton et  al. 1999). Thus, no maternal blood cells are found in the placenta during the first trimester, only a maternal plasma stream. This apparent lack of maternal blood cells in the intervillous space has been demonstrated by transcervical endoscopic examination of the intervillous space (Schaaps and Hustin 1988) as well as by Doppler ultrasound of the placenta (Jauniaux et al. 1991). However, there is a permanent flow of fluid through the intervillous space already

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in the first trimester. The plugs of the endoarterial trophoblasts do not allow blood cells to pass, but they do allow the passage of plasma, which then enters the intervillous space as a clear fluid. To this the secretion products of the uterine glands must be added, which are admixed with the plasma. This fluid, rich in lipids, nutrients and growth factors, is called uterine milk and is crucial for the nutrition and thus development of the placenta and embryo. Due to the absence of blood cells, only the oxygen physically dissolved in the plasma reaches the placenta during the first trimester. Thus, a physiologically low partial pressure of oxygen of 2 μm diameter were already described as being pinched off as syncytial knots from the syncytiotrophoblast at the end of the 19th century when they were discovered in the lungs of deceased pregnant women (Schmorl 1893). These fetal vesicles thus represent a further boundary or contact surface in the maternal organism. Microvesicles or ectosomes (diameter: approx. 100–1000 nm), which are released from the apical  syncytiotrophoblast membrane, and exosomes (diameter approx. 30–100 nm), which are released from intra-

cellular multivesicular structures, serve to communicate between the fetus or placenta and the maternal organism, including the immune system. The vesicles are composed of proteins, microRNA, RNA and DNA (Chamley et  al. 2014). Trophoblastic microvesicles express placental alkaline phosphatase (PLAP), among others, making them easily identifiable and distinguishable from vesicles of other origins (Gohner et al. 2015). In contrast, exosomes carry surface features from the cell interior of their cells of origin. It has been shown in  vitro that factors such as proteins or non-coding RNA produced in trophoblast cells can be transported via extracellular vesicles. When these vesicles are incubated with T lymphocytes, the trophoblast-derived RNA molecules are subsequently detectable in the lymphocytes and can influence the proliferation of target cells and thus the maternal immune system in pregnancy (Delorme-Axford et  al. 2013; Ospina-Prieto et al. 2016). The ­concentration as well as the composition of trophoblastic extracellular vesicles are often altered in pregnancy pathologies. In preeclampsia, their concentration in the blood of pregnant women is significantly increased (VanWijk et al. 2002). The fact that large amounts of extracellular vesicles enter the maternal circulation from the syncytiotrophoblast is unanimously accepted, but there are controversial results, especially about their composition in different diseases, largely stemming from the technical difficulties of isolation and analysis (Morales-Prieto et al. 2014).

2.4

Summary and Conclusion

In pregnancy, two allogeneic individuals live symbiotically together and their tissues, including the various immune cells, have immediate contact with each other. A num-

39 Immunology of the Fetomaternal Border

Anti-inflammatory

2

Pro-inflammatory Hormones

Progesterone PIBF hCG Prostaglandin E2 (PGE2)

Cytokines Th1 cytokines TNF, IFN-γ, IL-2, IL-12, IL-18

Th2 cytokines IL-4, IL-5, IL-6, IL-10, IL-13, TGFβ

Immune cells T helper cells Cytotoxic T cells M1 macrophages NK cells

γδ-T cells Treg cells uterine NK cells M2 macrophages

Trophoblast factors HLA class Ib HLA-G, HLA-E, HLA-F IDO Fas/FasLigand Galectin-1 extracellular vesicles

HLA class Ia HLA-A, HLA-B

..      Fig. 2.3  Schematic overview of important immunoregulatory factors at the fetomaternal interface. A relative predominance of anti-inflammatory factors

develops. Proinflammatory factors are partly completely absent or significantly reduced

ber of decidual and placental immunoregulatory cells and factors ensure the physiological course of pregnancy. In addition, the placenta releases factors into the maternal circulation that adjust her organism and immune system to pregnancy (. Fig.  2.3). It has been widely described that disturbances in the immunological balance can negatively affect fertility and pregnancy and may lead to disorders or disease. Their better understanding will lead to better diagnostic techniques as well as to new therapeutic approaches.

References

 

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Gordon S (2003) Alternative activation of macrophages. Nat Rev Immunol 3(1):23–35 Gordon S, Taylor PR (2005) Monocyte and macrophage heterogeneity. Nat Rev Immunol 5(12):953– 964 Guerin LR, Prins JR, Robertson SA (2009) Regulatory T-cells and immune tolerance in pregnancy: a new target for infertility treatment? Hum Reprod Update 15(5):517–535 Gustafsson C et al (2008) Gene expression profiling of human decidual macrophages: evidence for immunosuppressive phenotype. PLoS One 3(4):e2078 Heikkinen J et al (2003) Phenotypic characterization of human decidual macrophages. Clin Exp Immunol 131(3):498–505 Jabrane-Ferrat N, Siewiera J (2014) The up side of decidual natural killer cells: new developments in immunology of pregnancy. Immunology 141(4):490–497 Jones LA et  al (2010) Differential modulation of TLR3- and TLR4-mediated dendritic cell maturation and function by progesterone. J ­ Immunol 185(8):4525–4534 Kammerer U, von Wolff M, Markert UR (2004) Immunology of human endometrium. Immunobiology 209(7):569–574 Koc S et  al (2003) Enhancement of immunogenicity of Jeg3 cells by ectopic expression of HLA-­ A*0201 and CD80. Am J Reprod Immunol 50(3):243–253 Kopcow HD et  al (2008) T cell apoptosis at the maternal-­ fetal interface in early human pregnancy, involvement of galectin-1. Proc Natl Acad Sci U S A 105(47):18472–18477 Makrigiannakis A et al (2008) Fetomaternal immunotolerance. Am J Reprod Immunol 60(6):482–496 Martinez FO et al (2006) Transcriptional profiling of the human monocyte-to-macrophage differentiation and polarization: new molecules and patterns of gene expression. J Immunol 177(10):7303–7311 McIntire RH, Ganacias KG, Hunt JS (2008) Programming of human monocytes by the uteroplacental environment. Reprod Sci 15(5):437–447 Medawar PB (1953) Some immunological and endocrinological problems raised by the evolution of viviparity in vertebrates. Symp Soc Exp Biol 7:320–338 Mellor AL, Munn DH (2004) IDO expression by dendritic cells: tolerance and tryptophan catabolism. Nat Rev Immunol 4(10):762–774 Mincheva-Nilsson L (2003) Pregnancy and gamma/ delta T cells: taking on the hard questions. Reprod Biol Endocrinol 1:120 Moore KL, Persaud TVN (2007) Embryologie: Entwicklungsstadien, Frühentwicklung, Organogenese, Klinik, Bd 5. Elsevier & Urban & Fischer, München

41 Immunology of the Fetomaternal Border

Mor G (2008) Inflammation and pregnancy: the role of toll-like receptors in trophoblast-immune interaction. Ann N Y Acad Sci 1127:121–128 Mor G, Abrahams VM (2003) Potential role of macrophages as immunoregulators of pregnancy. Reprod Biol Endocrinol 1:119 Mor G, Cardenas I (2010) The immune system in pregnancy: a unique complexity. Am J Reprod Immunol 63(6):425–433 Morales-Prieto DM et al (2014) Elsevier trophoblast research award lecture: origin, evolution and future of placenta miRNAs. Placenta 35:39–45 Ospina-Prieto S et al (2016) MicroRNA-141 is upregulated in preeclamptic placentae and regulates trophoblast invasion and intercellular communication. Transl Res 172:61–72 Persson G et al (2017) HLA class Ib in pregnancy and pregnancy-related disorders. Immunogenetics 69(8–9):581–595 Piccinni MP et al (2000) Role of hormone-controlled Th1- and Th2-type cytokines in successful pregnancy. J Neuroimmunol 109(1):30–33 Poehlmann TG et al (2006) Inhibition of term decidual NK cell cytotoxicity by soluble HLA-G1. Am J Reprod Immunol 56(5–6):275–285 Porcheray F et  al (2005) Macrophage activation switching: an asset for the resolution of inflammation. Clin Exp Immunol 142(3):481–489 Redman CW, Sacks GP, Sargent IL (1999) Preeclampsia: an excessive maternal inflammatory response to pregnancy. Am J Obstet Gynecol 180(2 Pt 1):499–506 Reister F et al (2001) Macrophage-induced apoptosis limits endovascular trophoblast invasion in the uterine wall of preeclamptic women. Lab Invest 81(8):1143–1152 Reyes L, Wolfe B, Golos T (2017) Hofbauer cells: placental macrophages of fetal origin. Results Probl Cell Differ 62:45–60 Robinson DP, Klein SL (2012) Pregnancy and pregnancy-­ associated hormones alter immune responses and disease pathogenesis. Horm Behav 62(3):263–271 Saito S (2000) Cytokine network at the feto-maternal interface. J Reprod Immunol 47(2):87–103 Saito S, Sasaki Y, Sakai M (2005) CD4(+)CD25high regulatory T cells in human pregnancy. J Reprod Immunol 65(2):111–120 Santoni A, Carlino C, Gismondi A (2008) Uterine NK cell development, migration and function. Reprod Biomed Online 16(2):202–210 Schamberger S et  al (2013) Establishment of a one-­ sided ex vivo human placenta perfusion model to assess adhesion and invasion behavior of T cell leukemia cell lines. Leuk Lymphoma 54(8):1811– 1813

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Schmorl CG (1893) Pathologisch-anatomische Untersuchungen über Puerperal-Eklampsie. F. C, W Vogel, Leipzig Seshadri S, Sunkara SK (2014) Natural killer cells in female infertility and recurrent miscarriage: a systematic review and meta-analysis. Hum Reprod Update 20(3):429–438 Silini AR et  al (2017) Is immune modulation the mechanism underlying the beneficial effects of amniotic cells and their derivatives in regenerative medicine? Cell Transplant 26(4):531–539 Singh U et  al (2005) Immunological properties of human decidual macrophages—a possible role in intrauterine immunity. Reproduction 129(5):631– 637 Smith SD et  al (2009) Evidence for immune cell involvement in decidual spiral arteriole remodeling in early human pregnancy. Am J Pathol 174(5):1959–1971 Steinborn A et  al (2012) Pregnancy-associated diseases are characterized by the composition of the systemic regulatory T cell (Treg) pool with distinct subsets of Tregs. Clin Exp Immunol 167(1): 84–98 Svensson-Arvelund J et al (2014) The placenta in toxicology. Part II: systemic and local immune adaptations in pregnancy. Toxicol Pathol 42(2): 327–338 Svensson J et  al (2011) Macrophages at the fetal-­ maternal interface express markers of alternative activation and are induced by M-CSF and IL-10. J Immunol 187(7):3671–3682 Szekeres-Bartho J, Polgar B (2010) PIBF: the double edged sword. Pregnancy and tumor. Am J Reprod Immunol 64(2):77–86 Szekeres-Bartho J, Wegmann TG (1996) A progesterone-­dependent immunomodulatory protein alters the Th1/Th2 balance. J Reprod Immunol 31(1–2):81–95 Szekeres-Bartho J, Halasz M, Palkovics T (2009) Progesterone in pregnancy; receptor-ligand interaction and signaling pathways. J Reprod Immunol 83(1–2):60–64 Szekeres-Bartho J, Markert UR, Varla-Leftherioti M (2015) Immunology in reproduction. J Reprod Immunol 108:1 Szekeres-Bartho J et  al (2001) Progesterone as an immunomodulatory molecule. Int Immunopharmacol 1(6):1037–1048 Szekeres-Bartho J et al (2005) Progesterone-­dependent immunomodulation. Chem Immunol Allergy 89:118–125 Tao Y et al (2015) CD56(bright)CD25+ NK cells are preferentially recruited to the maternal/fetal interface in early human pregnancy. Cell Mol Immunol 12(1):77–86

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Terness P et  al (2007) Tolerance signaling molecules and pregnancy: IDO, galectins, and the renaissance of regulatory T cells. Am J Reprod Immunol 58(3):238–254 Trundley A, Moffett A (2004) Human uterine leukocytes and pregnancy. Tissue Antigens 63(1):1–12 Tuckerman E et al (2010) Uterine natural killer cells in peri-implantation endometrium from women with repeated implantation failure after IVF. J Reprod Immunol 87(1–2):60–66 Uckan D et al (1997) Trophoblasts express Fas ligand: a proposed mechanism for immune privilege in placenta and maternal invasion. Mol Hum Reprod 3(8):655–662

Vacca P, Mingari MC, Moretta L (2013) Natural killer cells in human pregnancy. J Reprod Immunol 97(1):14–19 VanWijk MJ et  al (2002) Microparticle subpopulations are increased in preeclampsia: possible involvement in vascular dysfunction? Am J Obstet Gynecol 187(2):450–456 Wegmann TG et  al (1993) Bidirectional cytokine interactions in the maternal-fetal relationship: is successful pregnancy a TH2 phenomenon? Immunol Today 14(7):353–356

43

Placental Morphology Berthold Huppertz and Thomas Stallmach Contents 3.1

Morphology of the Placenta – 44

3.1.1 3.1.2 3.1.3 3.1.4

I ntroduction – 44 Villous Development – 44 Development of Cell Columns for Trophoblast Invasion – 49 Structures on the Villous Surface – 53

3.2

Histopathology of the Placenta for Gynecologists – 54

3.2.1 3.2.2 3.2.3 3.2.4 3.2.5

I ntroduction – 54 First Trimester (Abortion) – 54 Second Trimester (Hydrops Fetalis, Infection and Inflammation) – 57 Third Trimester (Circulatory and Maturation Disorders) – 61 Postpartum Period – 67

3.3

Biobanking – 69

3.3.1 3.3.2 3.3.3

I ntroduction – 69 Variables Affecting the Composition of a Sample – 70 Collection or Biobank? – 71

Further Readings – 75

© Springer-Verlag GmbH Germany, part of Springer Nature 2023 B. Huppertz, E. Schleußner (eds.), The Placenta, https://doi.org/10.1007/978-3-662-66256-4_3

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44

3.1

B. Huppertz and T. Stallmach

Morphology of the Placenta

Berthold Huppertz

3

3.1.1

Introduction

3.1.2

Villous Development

First villi develop around the 5th week of pregnancy (post menstruation, p. m.) starting from the large trabeculae of the syncytiotrophoblast (Chap. 7 1). These trabeculae still range from the early chorionic plate to the basal plate and thus reach the maternal tissues. . Figure 3.1 shows such a trabecule at the 6th week of gestation with the first blood vessels in the placental mesenchyme and with syncytial sprouts leading to the development of the first mesenchymal villi. At this time, the intervillous space is still filled with blood plasma and glandular secretion products (uterine milk), maternal blood cells do not reach the placenta at this stage of pregnancy. The driving structures in the formation of the villous tree are the sprouts of the syncytiotrophoblast (. Fig.  3.2a). These syn 

The placenta is formed with the development of the trophoblast even before the implantation of the blastocyst. During the embryonic period and subsequently during the fetal period of pregnancy, the placenta must perform its multiple and changing functions, while at the same time undergoing massive morphological changes. This chapter section is devoted to the morphological features of the placenta over the duration of pregnancy and illustrates the features of the placenta in cross-­sectional images during the course of a healthy pregnancy.

 

 

Embryo (not in picture) Early chorionic plate with extraembryonic mesoderm

5th -6th week gestation

1

1

Trabecule Syncytial sprout

2

Placental mesenchyme First placental blood vessels 2

Villous trophoblast

3

Mesenchymal villus

3

Early intervillous space, not yet filled with maternal blood

Uterine tissue (not in picture)

..      Fig. 3.1  Villous development in the 5th/6th week of pregnancy. Starting from the early chorionic plate, a large trabecule can be seen which projects towards

the uterine tissue. The first villi develop from this trabecule by sprouting of the villous trophoblast

45 Placental Morphology

3

a 5th-6th week gestation Syncytial sprout Villous cytotrophoblast Formation of placental blood vessels Villous mesenchyme Early intervillous space, not yet filled with maternal blood 20 µm

Mesenchymal villus b 5th -6th week gestation Syncytial sprout Villous cytotrophoblast Early intervillous space, not yet filled with maternal blood Formation of placental blood vessels

Villous mesenchyme 20 µm

Hofbauer cell Mesenchymal villus

..      Fig. 3.2  a, b Villous development in the 5th/6th week of pregnancy. a A syncytial sprout indicates the development of a new villus. With the ingrowth of villous cytotrophoblasts, a primary villus is formed, which is then filled with connective tissue in its center

(secondary villus). b The continued growth of a syncytial sprout indicates the direction for the formation of a new villus. This sectional image shows the ingrowth of blood vessels into the new villus, which thus becomes a tertiary villus

cytial sprouts bulge into the intervillous space on the outside of the syncytiotrophoblast and define the areas where new villi will be formed. Here, the underlying cytotrophoblast proliferates, giving rise to the

first purely trophoblastic primary villi. Subsequently, villous mesenchyme pushes into these areas (secondary villus), followed by the development of blood vessels (tertiary villus). Thus, a complete tertiary villus

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is present, which continues to grow and develops more syncytial sprouts (. Fig. 3.2b), thus driving villous tree development. The mesenchymal villi are the first type of villus to develop and are characterized by a dense mesenchymal stroma with many mesenchymal cells, a few, still developing blood vessels and a thick layer of villous trophoblast (. Fig.  3.3). From these mesenchymal villi, the first additional type of villus develops from about the 8th week of pregnancy, the immature intermediate villus (. Fig.  3.3). This type of villus is the precursor of the stem villi and reaches significantly larger diameters than the mesenchymal villi. Immature intermediate villi have a very specific characteristic: the stromal channels. In these villi, the channels  

3

 

 

8th week gestation

run along the axis of the immature intermediate villi, are independent of blood vessels and are not lined by any epithelium or endothelium. Placental macrophages (Hofbauer cells) are frequently found in the stromal channels, which makes it easy to identify such cells in sectional images (. Fig. 3.3). In both types of villi, it is readily apparent that the villous trophoblast in the first trimester of pregnancy still consists of two complete layers: a complete layer of closely aligned villous cytotrophoblasts and a second layer consisting of the syncytiotrophoblast (. Fig. 3.3). Over the next few weeks of pregnancy, the composition of the types of villi does not change; this is primarily a time of further growth in the size of the placenta. Only starting at the 18th week of preg 

 

Mesenchymal villus Syncytiotrophoblast Villous cytotrophoblast Villous mesenchyme

50 µm

Immature intermediate villus Syncytiotrophoblast Villous cytotrophoblast Hofbauer cell Placental blood vessels Villous mesenchyme Stromal channels

50 µm

..      Fig. 3.3  Mesenchymal villus and immature intermediate villus at the 8th week of gestation. The images show a mesenchymal villus (top) and an immature intermediate villus (bottom). In addition to the typical histological features of these villi, the double layer of

villous trophoblast (syncytiotrophoblast and cytotrophoblast) can be seen in both types of villi as well as the stromal channels, which are only found in the immature intermediate villi

47 Placental Morphology

nancy, the immature intermediate villi begin to differentiate into stem villi. This occurs from the inside to the outside and is clearly visible in sectional images. . Figure  3.4 shows the differentiation of an immature intermediate villus into a stem villus. Due to a reorganization of the blood vessels, large central vessels develop in this villous type, around which extracellular matrix proteins (especially collagens) are now increasingly incorporated into the connective tissue. This process starts centrally around the vessels and continues outwards, so that at an advanced stage only marginal stromal channels are visible (. Fig. 3.4).  

 

3

After the stem villi have developed and are able to stabilize the villous trees, the remaining period of pregnancy is primarily dealing with increasing the exchange surface to allow sufficient nourishing of the fetus. This process is initiated with the development of the mature intermediate villi from about the 24th week of pregnancy. In histological sections, this type of villus is characterized by its elongated shape, its loose connective tissue and its many small blood vessel cross sections (. Fig. 3.5). At the end of the second trimester, it becomes obvious that the complete layer of villous cytotrophoblast is no longer present. Instead, isolated cytotrophoblasts  

Differentiating immature intermediate villus

20th week gestation

Stromal channels Central blood vessel Start of stromal fibrosing Differentiating stem villus 50 µm

Central blood vessels: artery (A) and vein (V) V

Progressive stromal fibrosing A

Remaining stromal channels

50 µm

..      Fig. 3.4  Differentiation of the immature intermediate villi into stem villi at the 20th week of gestation. While the immature intermediate villus in . Fig. 3.3 still shows the typical histological characteristics of this type of villus, it can be clearly seen that the immature intermediate villus in this figure (top left) is now differentiating towards a stem villus. The large vessel  

that develops in the center of the villus is striking. Further progressive differentiation leads to stem villi that are nearly complete (bottom right). Here the central vessels are surrounded by dense, fiber-rich connective tissue. Only at the margin, are there still remnants of loose connective tissue with stromal channels

48

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B. Huppertz and T. Stallmach

are found between the syncytiotrophoblast and the villous stroma. Although the villous cytotrophoblast proliferates significantly during gestation, the growth pressure of the villous stroma is so strong that it pushes the layer of the mononucleated cells apart, leaving only isolated cytotrophoblasts in the sectional image (. Fig. 3.5). The mature intermediate villi form the basis for the development of the terminal villi as the type of villus that provides the largest area for exchange between mother and child at term. Terminal villi develop  

24th week gestation

from about gestational week 27 and are characterized by a large number of capillaries as well as dilated capillaries (sinusoids) (. Fig. 3.6). This type of villus will make up the largest proportion of all types of villi at the end of pregnancy to ensure adequate growth of the fetus. Based on the above-mentioned processes of developmental changes in the types of villi throughout pregnancy, different villous sets result for the individual stages of development. These villous sets from the 15th to the 40th week of pregnancy are shown in . Fig. 3.7.  

 

Mature intermediate villi

Small blood vessels Loose stroma

Syncytiotrophoblast Villous cytotrophoblast 50 µm

Mature intermediate villi Syncytiotrophoblast Villous cytotrophoblast Small blood vessels Loose stroma 50 µm

..      Fig. 3.5  Mature intermediate villi at the 24th week of pregnancy. At the beginning of the second half of pregnancy, the mature intermediate villi develop as a prerequisite for the development of terminal villi.

This type of villus shows a distinct layer of the syncytiotrophoblast, under which only a few isolated cytotrophoblasts can be found

3

49 Placental Morphology

31th week gestation

Terminal villi/ Mature intermediate villi

37th week gestation

Capillaries Sinusoids

Syncytiotrophoblast Loose stroma Villous cytotrophoblast

20 µm

..      Fig. 3.6  Terminal villi and mature intermediate villi at the 31st and 37th week of gestation. These two types of villi are hardly distinguishable in diameter.

3.1.3

Development of Cell Columns for Trophoblast Invasion

Trophoblast invasion by the extravillous trophoblast starts in the 5th week of pregnancy (p. m.) and continues throughout the entire duration of pregnancy. Initially, starting from a small placenta, a large number of invasive cells penetrate the maternal tissues of the uterus. As pregnancy progresses, the placenta expands and trophoblast cells can invade from a greater number of entry sites. The trophoblast cell columns as a source of invading cells thin out over the course of pregnancy, so that by term these cell columns are mostly completely depleted. In the first trimester of pregnancy, multilayered cell columns are found that extend

20 µm

Here, the presence of sinusoids must serve as a differential diagnostic feature, which distinguishes terminal villi from mature intermediate villi

far beyond the boundary of the decidual tissue into the placenta, where they are surrounded by maternal blood plasma. This picture can be seen in the 5th–6th (. Fig. 3.8a), as well as in the 8th week of pregnancy (. Fig. 3.8b). In the second trimester, the cell columns remain intact and a large number of extravillous trophoblasts find their way into the maternal tissue of the uterus (. Fig.  3.9). The base of the anchoring villi widens, so that the cell columns also become wider and allow a better hold for anchoring the placenta to the uterine wall. In the last trimester of pregnancy many cell columns are used up. Now the anchoring villi are firmly anchored to the basal plate, while only remnants of cell columns are visible (. Fig. 3.10). Through the secre 

 

 

 

50

B. Huppertz and T. Stallmach

IIV

IIV

ST

IIV

3 15th week

24th week

20th week

ST

ST 28th week

31st week

37th week

ST

40th week ..      Fig. 3.7  Overview images of placentas between the 15th and 40th week of pregnancy. Over the course of pregnancy, it becomes obvious that the villi become smaller in caliber, but at the same time increase in number. This leads to a significant increase in surface area, which can be used for nutrient and gas exchange.

It can also be seen that the large immature intermediate villi are very prominent in the first half of gestation, but are hardly found with the emergence of the mature intermediate villi. IIV immature intermediate villus; SV stem villus

..      Fig. 3.8  a, b Cell columns in the first trimester. In the basal region of an anchoring villus the trophoblastic cell columns are found, from which the extravillous trophoblasts invade the uterine tissues of the mother. In the proximal portion of the cell columns (a, asterisk) are the cells that are still actively proliferating. Proliferative pressure pushes subsequent generations of daughter cells toward the uterus. These cells then separate from each other and start their invasion

into the decidua (b). The cells in the area of proliferation are clearly distinguishable morphologically from their daughter cells. While the proliferative cells are still small with dense and clearly stainable cytoplasm, the post-proliferative daughter cells are much larger and store glycogen, which is washed out during embedding. Hence, these cells appear pale and empty in a section (a, arrowheads)

3

51 Placental Morphology

a

Anchoring villus

Anchoring villus with cell column

5th-6th week

Ce

ll c

Villous stroma

olu

mn

Syncytiotrophoblast *

Villous cytotrophoblast

∆

Extravillous cytotrophoblast

∆ Decidua

∆ 50 µm

b

Anchoring villi with cell columns

8th week

Villous stroma Syncytiotrophoblast

Anchoring villus

Villous cytotrophoblast

Anchoring villus

Extravillous cytotrophoblast

Cell column

Decidua 50 µm

52

B. Huppertz and T. Stallmach

Anchoring villi with cell columns

3

Villous stroma

15th week Anchoring villus Cell column

Syncytiotrophoblast Villous cytotrophoblast Anchoring villus

Extravillous cytotrophoblast

Cell column Decidua 100 µm

..      Fig. 3.9  Cell columns in the second trimester. In the second trimester the cell columns flatten and are no longer as high as in the first trimester. However,

large amounts of extravillous trophoblasts continue to be formed, which invade the uterine wall and open uterine luminal structures

Anchoring villus with cell column

28th week

Villous stroma Syncytiotrophoblast Villous cytotrophoblast

Extravillous cytotrophoblast

*

Adhesive pad

Remainder of a cell column Decidua 100 µm

..      Fig. 3.10  Cell column in the third trimester. Cell columns are still found even in the third trimester. However, they can no longer perform their function as a source for the production of the supply of extravillous trophoblasts, as the proliferative part of the cell columns has been used up. The anchoring villi now

extend into the decidua, are fibrosing in their center (asterisk) and are surrounded on their basal side by extravillous trophoblasts that have secreted large amounts of matrix-type fibrinoid. This allows the villous portion of the placenta to be well fixed to the basal plate

53 Placental Morphology

tion of matrix-type fibrinoid, the extravillous trophoblasts further solidify the area of the basal plate. 3.1.4

Structures on the Villous Surface

Especially in the mature placenta, different structures on the villous surface are often used to differentiate pathological from nor-

3

mal pregnancies (. Fig.  3.11). On the one hand, there are protrusions of the syncytiotrophoblast such as the syncytial sprouts for villous development or the syncytial knots for the release of apoptotic nuclei. On the other hand, there are also changes in the three-dimensional structure of the villous trees, which result in changes in the sectional images and can be erroneously interpreted as protrusions. Thus, these structures are not always correctly assigned.  

41st week of gestation

..      Fig. 3.11  Structures on the villous surface at the 41st week of pregnancy. In a mature placenta, corpuscular structures can be visualized on the villous surface. These are readily used to differentiate pathologic from normal pregnancies without distinguishing between these structures. Typically, at least three types of these structures can be distinguished: Outgrowths of the syncytiotrophoblast for villous development (syncytial sprouts), outgrowths of the syncytiotro-

phoblast for the release of apoptotic material (syncytial knots), and flat sections through the villous trees, which are difficult to distinguish from outgrowths. Since this is a placenta from the 41st week of gestation, there are two possibilities for the structures shown: syncytial knots or flat sections through the syncytiotrophoblast. Only a three-­dimensional reconstruction of serial sections can clearly assign the true nature of such corpuscular structures

54

3.2

3

B. Huppertz and T. Stallmach

Histopathology of the Placenta for Gynecologists

Thomas Stallmach 3.2.1

Introduction

The placenta is not comparable to any other organ. As it grows and develops its final architecture (maturation), it must perform a range of functions that will later be distributed among several organs. However, it only fulfils the requirements for nine months, and in some ways you can see it doing so. A tissue sample of the body organs of a young healthy person is usually immaculate under the microscope, while tissues from a term placenta without any complications always show read off pathological changes under the microscope. This makes the histopathological examination of a placenta a special task, in which the quantity of a change often plays a greater role than its quality. Thus, perivillous fibrin deposits in small quantity are apparently physiological, to be understood as the most favorable variant for repair of tissue defects in view of the remaining life of the organ. Above a certain amount of deposits and in the presence of further changes, however, the diagnosis of a maternal circulatory disorder arises, as is frequently observed in the context of preeclampsia. Unlike in the case of bodily organs, every examination of the placenta raises the question of whether a morphological finding is significant at all. If so, two aspects arise: 1. Were the findings so pronounced that a supply restriction or threat existed intrauterine or peripartum? This can be read off from the condition of the newborn if the nutritive function of the placenta was restricted, but is difficult to assess if the oxygen supply to the child was restricted.

2. Does the finding point to an underlying condition in the mother or child that may still be significant postpartum? With largely perfected obstetrics, the vast majority of pregnant women will give birth to a healthy child, freed at this moment from the latent threat of a not always perfect short-lived multifunctional organ. In the following, placental pathology is presented arranged according to gestational age and disease groups, with a sentence at the end on the usefulness or necessity of postpartum examination of the organ appended to each group. 3.2.2

First Trimester (Abortion)

Human pregnancies are characterized by a relatively high rate of spontaneous pregnancy losses; this amounts to approx. 10% overall. After an abortion, the risk of a further ­abortion increases to 15%, and thereafter to 25%. The expelled tissue is usually sent for pathological examination to exclude malignant transformation of the trophoblastic tissue and to obtain an indication of the cause of the abortion.

Abortion with and Without Developmental Aberration Apart from the exclusion of malignant transformation, the morphological examination of abortion material by a pathologist actually permits the clarification of only one question: Is the morphology of the placenta, in particular of the chorionic villi, so abnormal that a primary defect of the embryo must be assumed with great probability as the cause of the abortion? In the only two-­dimensional sectional picture, the root-like branches of the chorionic villi reveal a certain harmony in an undisturbed pregnancy. The stroma of the chorionic villi is ­ uniformly loose, equipped with fetal

55 Placental Morphology

blood vessels. The trophoblastic cover appears predominantly bilayered with polar broadenings interpreted as growth zones (. Fig. 3.12a). If the pregnancy is disturbed from the embryonic side, all three parameters may be disturbed. The architecture of the villous branching is irregular, the stroma is poor of cells and very loose, often in combination with villi whose stroma appears compacted, the trophoblast is partly atrophic, partly irregularly proliferating (. Fig.  3.12b). Some of the changes may be pretended by regressive changes, which are almost the  

 

rule in the setting of a “missed abortion.” If there is clear pathology in at least two of the three criteria (architecture, stroma, trophoblast), the diagnosis is “developmental aberration of chorionic villi”. From this, a “primary embryonic cause of the abortion” can be deduced, which—as analyses of tissue from abortion material have shown—is usually based on a chromosomal aberration. The tissue of the placental basal plate and decidua often shows a band-like inflammation, which has set in as a result of the death of the embryo/fetus to demark the tissue.

a

b

c

d

..      Fig. 3.12  a Regular morphology of the chorionic villi in the first trimester: regular branching pattern, regular nature of the villous stroma and two-layered trophoblast. b Aberration of the chorionic villi: irregular branching pattern, loose and dense stroma in alternation, atrophic trophoblast sections and trophoblast proliferation patterns. c Partial hydatidiform mole: Two morphologically distinct populations of chorionic villi, (I) nearly regular small villous caliber,

3

(II) villous vesicle (fluid accumulation without stroma) with trophoblast proliferation, usually not covering the whole circumference of the villi and not showing severe atypia. d Complete hydatidiform mole: only vesicularly transformed chorionic villi (no normal villous population) with abundant trophoblast proliferation that often captures the entire circumference of the villi and shows distinct atypia (arrow). (H&E, 100×)

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If a developmental aberration cannot be determined morphologically with the necessary certainty, the diagnosis can only state the absence of this finding. In some of these cases, necrotic chorionic villi surrounded by fibrin and hemorrhages of different ages are seen in the decidua, which is occasionally summarized as an “embryonic bed disorder”. It remains completely open whether this is a primarily anatomical, hormonal or immunological problem, by which a subsequent pregnancy could also be affected, or a statistical event in the incipient remodeling of the maternal circulation in the region of the placenta.

Question About Hydatidiform Mole Partial and complete hydatidiform moles also fall under the term “developmental aberration”. However, the morphological picture is so typical that the chromosomal aberration behind it can be inferred immediately. In the case of a complete hydatidiform mole, this results in a specific risk profile. In the case of partial and complete hydatidiform mole, the term is derived from the vesicular enlargement of the chorionic villi. Strictly to be distinguished from this is the “blighted ovum”, in which an empty amniotic sac is seen as a vesicle. The absence of an embryo in a blighted ovum” is almost always an expression of a “developmental aberration” and will be accompanied by the above-mentioned morphological criteria of the chorionic villi. Etiologically, this is due to a multitude of spontaneously occurring chromosomal aberrations, but not to the specific chromosomal findings of hydatidiform moles. Etiologically, the partial hydatidiform mole is based on a triploidy, i.e. in each cell of the embryo there is a triple set of chromosomes (karyotype 69 XXX, 69 XXY or 69 XYY). If two sets of chromosomes originate from the mother, the result is a dystrophic pregnancy with very small embryos/

fetuses displaying characteristic anomalies. If two sets of chromosomes are of paternal origin, the result is a partial mole. An embryo is not formed in this case. Morphologically, chorionic villi fall into two distinct populations: largely inconspicuous chorionic villi contrast with vesicularly distended chorionic villi with irregular trophoblast. This gives rise to proliferation sites that are not polar (see above) but grow circumferentially in several directions without, however, showing any appreciable atypia (. Fig. 3.12c). The complete mole is etiologically based on a zygote with two paternal chromosome sets without maternal chromosomal material. Again, no embryo is formed. All chorionic villi are vesicularly degenerated, do not show any blood vessels and are circumferentially covered by an exuberant trophoblastic proliferation within which atypical nuclei can be found (. Fig. 3.12d). The risk of a complete hydatidiform mole to progress into a choriocarcinoma is variably reported; both the frequency of complete moles and the risk of their malignant transforamtion appear to depend on ethnic factors. With an average risk of 1:50, a longer period of contraception is recommended to be able to detect the development of choriocarcinoma at an early stage by means of regular checks of βhCG.  

 

Answers and Evidence from Examination of Abortion Material The suspicion of a hydatidiform mole arises at the latest from the 10th week of pregnancy (ultrasound). In the case of abortions obtained earlier, the diagnosis of a hydatidiform mole can occasionally be made without any previous clinical suspicion. At the time of removal of a hydatidiform mole, it is unlikely that a pre-existing choriocarcinoma is present. Only complete hydatidiform moles, but not partial moles,

57 Placental Morphology

require a prolonged period of βhCG monitoring to rule out a choriocarcinoma. The risk of aberration after a partial mole is not different from the basal risk of aberration of trophoblastic tissue inherent in all pregnancies, this is reported to be 1:25,000. Thus, while the detection of specific findings (hydatidiform mole, congenital storage diseases already manifesting in the trophoblast) are rarities, the morphological evaluation of any abortion should include the statement: “developmental aberration present/not present/no definite statement possible at this point”. Face to face with the patient, the findings can be interpreted as follows. In the majority of cases, the abnormality refers to a chromosomal aberration (usually trisomy) in the embryo. In the vast majority of cases, such an aberration has occurred by chance during the development of the oocyte at the time of maturation. It is not necessary to know which aberrations are present. What is decisive is that the maternal organism (here the choice of words is a little delicate) has recognized the problem and the pregnancy has been terminated. It is not a negative omen for further pregnancies. This interpretation is not entirely without limitation. If the pregnant woman or her partner (!) is the carrier of a balanced chromosomal translocation, there is a systematic risk. In this case, one third of all further pregnancies will again be miscarriages. Of the delivered, (phenotypically) healthy children, half will also have the translocation. However, the statistically expected frequency of a pregnancy in which one of the partners is a translocation carrier is so low that this alone is not an indication for chromosomal testing of the parents after an abortion with a developmental aberration. In the case of abortions without evidence of a developmental aberration, the counselling situation seems less clear, since the cause cannot be clearly stated as being on the side of the embryo.

3.2.3

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Second Trimester (Hydrops Fetalis, Infection and Inflammation)

Hydrops of Fetus and Placenta Subcutaneous edema and effusions in the body cavities are called hydrops fetalis. In most cases the placenta is also hydropic and consequently pale and enlarged. The cause is a heterogeneous spectrum of diseases, led by various forms of fetal anemia. Hydrops may also be a consequence of congenital heart and lung malformations or an accompanying phenomenon of congenital tumors. In about 15% of cases there is a chromosomal aberration (e.g. monosomy X). Up to 30% of all cases remain unexplained.

I mmunological Hydrops in Blood Group Incompatibility Maternal antibodies directed against cells of fetal erythropoiesis destroy these cells and lead to fetal anemia. Prior to the recognition of the correlations and the development of effective therapy and prophylaxis (from 1969 onwards anti-D prophylaxis), these were mostly antibodies against the rhesus factor with prior sensitization of the mother, often through a previous pregnancy or abortion. In migrating populations that were previously poorly provided with medical care, the rhesus problem can now be increasingly observed again. In addition, sensitivities to other blood group antigens can rarely cause the same clinical picture. Hydrops of the placenta leads to pale and fragile tissue. This and the increase in size of the organ can occur rapidly analogous to the rapid development (but also possibility of improvement) of pulmonary edema in adults. The reason lies in the intrauterine circulatory situation. Both heart chambers work in parallel, the lung lies in a shunt. If increased circulatory work (due to anemia) leads to cardiac enlargement, muscular

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insufficiency or even insufficiency of the AV-­ valves, this leaves the fetal lungs largely unaffected, but leads to congestion and development of edema in the systemic circulation. Microscopically, the edema of the chorionic villi is also striking, in addition to an increased number of Hofbauer cells. Numerous nucleated erythrocytes are found in the fetal blood vessels—otherwise hardly detectable in the second trimester (. Fig. 3.13a). If the condition persists for longer, the chorionic villi also appear more  

immature with regard to their trophoblast covering compared to the gestational age.

 ydrops as A Result of Infection H (Fifth Disease, Erythema infectiosum) The destruction of cells of fetal erythropoiesis can also be the work of a virus. Parvovirus B19 is the causative agent of the generally little-known fifth disease. About 50% of women of childbearing age are immune. In the context of small epidemics, b

a

* * * *

c

..      Fig. 3.13  a Hydrops in infection by parvovirus B19: The chorionic villi show increased fluid retention (= edema), which becomes obvious by lifted trophoblastic epithelium (*) in histological sections. In the fetal blood vessels there is a large number of nucleated erythrocytes, which have passed from the sites of fetal hematopoiesis into the circulation (arrows). This finding also arises as a consequence of destruction of erythrocytes in rhesus incompatibility. In addition, large erythropoietic cells with virus-typical nuclear inclusions are seen in the fetal circulation (arrowhead). b Acute villitis in maternal sepsis: Dense infiltrate of

d

neutrophilic granulocytes within the chorionic villi. In a silver stain Campylobacter jejuni could be visualized. c Focal non-specific villitis in premature birth in the 30th week of gestation without evidence of infection: small foci of fibrin-clotted chorionic villi with loose mononuclear infiltrate. a–c H&E, 400×. d Early stage chorioamnionitis: maternal neutrophilic granulocytes as a dense fringe in the intervillous space below the chorionic plate (arrows). Fetal inflammatory cells have emerged from fetal blood vessels in the chorionic plate close to the amnion (arrowhead). (H&E, 200×)

59 Placental Morphology

seroconversion occurs in 3–7% of pregnancies. Symptoms occur in only 30% of infected pregnant women (transient anemia and skin rash); of the fetuses, only a small proportion becomes transplacentally infected. Within the fetal organism, apoptosis is induced by the virus in nucleated precursor cells of erythropoiesis; the resulting fetal anemia leads to hydrops due to circulatory mechanisms as in hydrops of immunological origin (. Fig. 3.13a). Since the virus is apparently capable of inducing fetal anemia for only a short time, it does not damage any other organs, and soon becomes undetectable. A rapid and marked improvement of the hydrops can be obtained by transfusion of blood to the fetus.  

Placentitis Placentitis (synonym “villitis”) describes the morphological picture of inflammation in and around chorionic villi. Acute placentitis (. Fig. 3.13b) is caused by a group of viral and bacterial pathogens which spread hematogenously to the placental villi during maternal infection. Entry into the fetal tissues leads to inflammation, which can result in typical defects or malformations (e.g. rubella virus, varicella-zoster virus, cytomegalovirus, listeria and Treponema pallidum) or, via fulminant sepsis, to the death of the fetus (e.g. streptococci, staphylococci, campylobacter). If the morphological examination of the placenta reveals an inflammation of the chorionic villi without evidence of pathogens and usually without a clearly defined clinical picture in the fetus, this is referred to as chronic placentitis. The majority of these cases do not receive satisfactory etiological clarification.  

(Acute) Inflammation of the Chorionic Villi with Evidence of Pathogens 55 Rubella virus (German measles): An initial infection of the mother during pregnancy also leads to viremia in the fetus.

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The growth of fetal cells is temporarily slowed down by the infection. In the context of an inflammatory tissue reaction, malformations occur. The placenta shows a necrotizing inflammation in the chorionic villi, typically involving the fetal vessels (endarteritis/endangiopathia obliterans). 55 Varicella zoster virus (chickenpox): The rare initial infection of the mother during pregnancy (especially in the period 8-20 weeks of gestation) leads to viremia in about 2% of fetuses, in which nerve cells in particular are attacked. Tissue atrophy occurs in the supply area of the associated nerve tracts and even mutilation of the extremities. The placenta shows only focal inflammation with necrosis and cell changes, which are suspicious for the presence of a viral infection. 55 Cytomegalovirus (CMV): Not only the initial infection of the pregnant woman, but also reactivation during pregnancy with existing maternal immunity can lead to severe fetal damage. In the case of fetal survival, this results in particular in central nervous defects. In cases of fetal death (30% of cases), abundant characteristically altered virus-infected cells (owl eye cells) are found in the tissues. In the associated placenta, the detection of owl eye cells is a rarity; usually only a (non-specific) lymphoplasmacytic inflammatory infiltrate is observed. 55 Campylobacter jejuni: Campylobacter is ingested from infected food (milk, poultry meat) and leads to acute bacterial colitis in the pregnant woman, which rapidly resolves. Apparently due to the altered immune status during pregnancies, maternal sepsis may develop and pass hematogenously to the fetus. While fetal sepsis is always fatal with Campylobacter, maternal deaths are rare. The chorionic villi show villitis with neutro-

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phil granulocytes (. Fig.  3.13b), and special staining can be used to see coiled bacilla. 55 Listeria monocytogenes: Listeria are ubiquitous, are ingested through infected food and rarely lead to sepsis. In case of altered immunity and high infectious dose, sepsis may result in mother and fetus. Abscessing inflammation occurs in fetal tissues, and fetal lethality is 50% with residual central nervous damage in survivors. The chorionic villi show micro-abscesses with histiocytic cells, often involving the fetal membranes (Section “Chorioamnionitis”). 55 Toxoplasma gondii: Toxoplasmosis is a protozoa-caused infectious disease that is widespread in animals. Infection during pregnancy through infected food or contact with acutely diseased domestic animals can lead to hematogenous transplacental infection of the fetus. Intrauterine growth restriction occurs, preterm birth with psychomotor retardation usually results, and fetal death is rare. The placenta shows no inflammatory infiltrates, but intracellular pseudocysts in the amniotic epithelium, in which the pathogen lies in the form of the so-­ called tachyzoites.  

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(Chronic) Inflammation of the Chorionic Villi Without Evidence of Pathogens In 5–10% of the patients examined (frequency depends on the indication for examination of a placenta), areas of fibrin-clotted chorionic villi (so-called Gitterinfarcts) with infiltrates of lymphocytes and macrophages, occasionally also plasmacytoid cells and giant cells are found (. Fig.  3.13c). Pathogen detection is not successful. Among the children born, there is a somewhat high incidence of premature births and “smallfor-date children”; while there is no evidence  

of an infection. A placenta during CMVinfection may show such a relatively low morphological finding, especially without specific findings (owl eye cells; see above), that only after exclusion of such an infection in newborns the diagnosis of a focal unspecific villitis (VUE, “villitis of unknown etiology”) can be definitely made. It is most likely to be the mitigation of immunological phenomena at the fetomaternal border or the reaction to a primary intervillous circulatory disturbance.

Chorioamnionitis (Amniotic Infection Syndrome) Physiologically, the lower genital tract is heavily colonized with bacteria, while the amniotic fluid is sterile. If bacterial colonization of the amniotic fluid does occur, both the maternal and the fetal organism react. Maternal granulocytes first infiltrate the subchorial fibrin (stage I; . Fig.  3.13d), then the covering tissue (stage II), and finally lead to necrosis of the amniotic epithelium (stage III). Fetal granulocytes are seen first within the blood vessels of the chorionic plate (stage I; . Fig. 3.13d), then in the wall of the umbilical artery (stage II), finally followed by necrotizing inflammation of the umbilical stroma (stage III). The most frequent consequence of the developing chorioamnionitis is premature birth. Intrauterine physiologic aspiration of amniotic fluid brings abundant inflammatory cells (fetal and maternal granulocytes) into the immature fetal lung tissue, which may be formally termed “intrauterine pneumonia.” Surprisingly, only a small proportion of preterm infants show signs of infection. The aspirated inflammatory infiltrate is clinically associated with a clustered occurrence of “wet lungs”, usually antibiotic treatment is not necessary. However, neonatal sepsis is more frequent in the collective “preterm infants with chorioamnionitis” compared to an unselected collective of born infants.  

 

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Umbilical Cord Infection An inflammatory infiltrate in the umbilical cord is usually part of the morphological picture of chorioamnionitis and correlates with its temporal course. Frequently, the fetal granulocytes are seen only in the wall of an umbilical artery and are thereby sectorially directed against the surface of the umbilical cord (stage II). Rarely, necrotizing inflammation in Whartonʼs jelly leads to necrosis (stage III of the fetal response) or, in the later stages, to calcifications that are already visible to the naked eye. This needs to be distinguished from a fungal infection (Candida), in which macroscopic yellowish spots can be seen on the surface of the umbilical cord, microscopically accompanied by granulocytes without reference to the umbilical cord vessels.

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fetus by autopsy is not permitted, some cases can be satisfactorily clarified by examination of the placenta. It should be noted, however, that in the case of diseases (e.g. CMV), which are rapidly apparent on microscopic examination of the tissues of deceased infants, only discrete focal findings may be obtained in the placenta. 3.2.4

Third Trimester (Circulatory and Maturation Disorders)

In the third trimester, the placenta must provide the fetus, which is in principle also viable outside the uterus, with the basis for a further 5- to 6-fold increase in body weight. This process can be restricted by disturbances of the blood circulation or the maturation of the chorionic villi. If the Answers and Evidence pathological process particularly affects from Examination of Placentas fetal nutrition, intrauterine growth restricin Preterm Delivery tion occurs. If this is a warning sign, premaThe diseases treated here as disorders of the ture delivery can be considered with the help second trimester may also extend to the of other diagnostic aids (e.g. Doppler ultrathird trimester, and some of the entities that sound of fetal and placental blood flow). If will be discussed there (e.g. preeclampsia) the pathological process affects fetal oxyoften already affect the second trimester. In genation in particular, no warning signs are view of the overall low number of births in seen. The sad consequence is sudden intrathe second trimester and the usually serious uterine fetal death, which is responsible for pathology involved, the recommendation about 25% of perinatal mortality in regions could be made to subject every placenta of well-developed obstetric care. Individual from this period to a pathological-­ children can be saved if reduced oxygenanatomical examination. With a strict focus ation (restricted placental reserve capacity) on individual diagnosis, one can leave all is revealed by an “accidentally” performed placentas without examination as an excep- oxytocin stress test. tion to this rule if they originate from circumstances that have already been clearly Maternal Circulatory Disorder diagnosed (e.g. fulminant infection with an The fetal chorionic villi are circulated identified pathogen). Placentas resulting around by maternal blood. Regional arrest from interruptions in this period, whether of maternal blood circulation in the intervildue to chromosomal aberration or prena- lous space results in spatially limited necrotally identified malformations, also do not sis of chorionic villi. Larger, approximately require examination. In the case of a spherical necrotic areas result from ischemia deceased child without a clear diagnosis, the of a fetal flow path unit (cotyledonary examination of the fetus and placenta is of infarction; . Fig.  3.14a); smaller ischemic primary importance. If examination of the districts, variable in shape, are termed  

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a

b

Infarct

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ZZM

..      Fig. 3.14  a (Maternal) cotyledonary infarction: Large roundish limited district of intervillous fibrin and devitalized chorionic villi (infarction). The unaffected chorionic villi show preterm maturation to terminal villi. In the two-dimensional image of the chorionic villi many small calibers can be seen, while the growth compartment of the placenta in the form of the intermediate villi is reduced in quantity. The caliber gap between the stem villi and the terminal

villi leads to the concept of intermediate villous deficiency (IVD). (H&E, 30×). b (Fetal) Endangiopathia obliterans: The blood vessels of a stem villus show an extensive lumen loss due to connective tissue proliferation sites, which are repeatedly interspersed by erythrocyte extravasations (arrow); in places recanalization is indicated (arrowhead). Neighboring chorionic villi show reduced to absent vascularization. (H&E, 150×)

Gitterinfarcts. In vivo, the maternal space between the chorionic villi (intervillous space) is of capillary width only. If the chorionic villi are forced apart by maternal blood clotted in the intervillous space, this is referred to as an intervillous thrombus.

phoblast) should have infiltrated the muscular walls of the maternal spiral arteries and remodeled them into wide muscle-free sections. If the activity of the extravillous trophoblast was too low, the inflow of maternal blood to the intervillous space is reduced with the consequences of decreased placental growth and the occurrence of infarcts (preferably Gitterinfarcts). Examination of the placental basal plate reveals a typical finding: maternal spiral arteries in which the intramural trophoblast cells are absent and which instead still have smooth muscle cells, interspersed in places with foam cells (acute atherosis).

Preeclampsia Preeclampsia and HELLP syndrome (“hemolysis, elevated liver enzymes, low platelet counts”) are clinically and laboratory-­chemically defined disease states which, from an etiological point of view, conceal different disease processes. In severe cases, which become symptomatic as early as the second trimester and are associated with fetal growth restriction, examination of the placenta (typically in preterm birth, 25–28 weeks of gestation) reveals a defective remodeling of the maternal blood circulation to the intervillous space. Fetal-derived trophoblastic cell invasion (extravillous tro-

Premature Placental Abruption Premature placental abruption is a clinical diagnosis caused by a retroplacental hematoma. Its development may be favored by systemic (hypertension) and local factors (pathology of the basal plate in preeclamp-

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sia). Examination of the placenta reveals an indentation of the basal plate with firmly adherent blood coagula. Further examination may reveal regional (small) infarcts and/or intervillous thrombi. These signs of a not only fresh intervillous circulatory disturbance are absent if the premature abruption was fulminant (e.g. due to abdominal trauma).

Fetal Circulatory Disorder A placental circulatory disturbance on the fetal side is morphologically most likely to be recognized by the blood vessels in the section of the stem villi (endangiopathia obliterans). Unfortunately, the finding is not specific as to etiology, as it may occur in the setting of an infection (e.g., rubella infection) or as a reaction to an upstream flow obstruction (umbilical cord complication) and, to some extent, even as an irrelevant secondary finding following fetal death from another cause.

Endangiopathy Obliterans If the placenta is macroscopically inconspicuous in intrauterine fetal death, the microscopic findings in the fetal vascular system often point the way. Fluctuating blood pressure and flow in the fetal circulation are a stimulus that induces endothelia to proliferate. Vascular cross-sections in the stem villi show restriction of the lumen by cell proliferation, sometimes with fragmented erythrocytes (. Fig.  3.14b); in later stages, connective tissue occlusions with multiple small capillary lumina (recanalization) are found. If this endangiopathy obliterans (“fetal thrombotic vasculopathy”) is accompanied by inflammatory infiltrates, an infection must be sought (e.g. rubella, cytomegaly). The findings of an endangiopathy obliterans must be clearly recognizable and must be identified in numerous vessel cross sections. If the vessel wall is only edematous with dehiscence of the cells and presumed lumen restriction, this may be a  

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phenomenon after an intrauterine fetal death, which has already occurred some time ago and which cannot be attributed to a vascular pathology.

Hereditary Thrombophilias Fetuses that have received several thrombophilic hereditary factors from both parents (e.g. factor V Leiden, protein S deficiency) may already show intrauterine consequences of this multiple heterozygous condition within the coagulation cascade. The clear endangiopathy obliterans that develops early in this process is typically accompanied by a circulatory disturbance also in the maternal circulation (in the form of Gitterinfarcts). A coagulation-physiological clarification of the parents (today mostly in the form of genetic tests) is only indicated if signs of thrombophilic diseases can be elicited in the family history.

Umbilical Cord Complications A true, tightly tightened knot in the umbilical cord is plausible as a cause of fetal circulatory disturbance. Obviously, also umbilical cord wrappings, in consequence of which a section of the umbilical cord is put under strong tension by fetal movements (between the base of the umbilical cord and the place of the wrapping) may lead to fluctuations of the blood flow. A clearly recognizable endangiopathy obliterans is then to be regarded as pathogenic; the finding of umbilical cord looping (in a dead child) is thereby recognized as pathogenetically relevant.

Maturation Disorder Fetal blood flows through the umbilical cord and is distributed over the chorionic plate of the placenta, from where it finally circulates through the chorionic villi, which are densely packed in a “pot” (intervillous space) formed by the mother. The chorionic villi can be divided morphologically into three segments. The stem villi have blood vessels with muscular walls and a dense

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stroma; they distribute blood to the intermediate villi. These are considered the growth compartment in the villous trees; blood vessels have a thin wall and the stroma is loose. The terminal villi are the actual functional compartment for exchange (. Fig.  3.15a); it is here that the most impressive changes in morphology are found during pregnancy. The term maturation is used to describe  

qualitative and quantitative changes in the three types of villi during pregnancy. The term “maturation disorder” refers to clearly recognizable deviations from a morphological ideal type of villous differentiation (maturation) related to the respective gestational age. Pathogenetically, the maturation disorder may be an adaptive phenomenon (e.g. “intermediate villus deficiency” in the case

a

b

c

d

..      Fig. 3.15  a Normal villous maturity at term: From the 34th week at the latest, mature terminal villi dominate the picture quantitatively: The small villous cross-sections are occupied to approx. 30% by blood vessels, whose sinusoidally dilated lumina are only separated from the maternal intervillous space by a thin cytoplasmic membrane (syncytio-capillary membranes, arrows). b Chorangiosis type II: Morphological continuum of intermediate and terminal villi with enlarged cross-sections compared to the normal state and a significantly higher proportion of cross sectional area allotted to blood vessels. Sinusoidal and syncytio-capillary membranes are abundantly formed. c Retardation of villous maturation in diabetes: the morphological development of the placenta towards

term stays behind “concordantly”: both the branching pattern and the extent of vascularization of the chorionic villi appear to correspond to a much earlier gestational age. Macroscopically, the placenta is usually large, the sectional surface is too pale for the gestational age. d Terminal villus deficiency: the chorionic villi show a normal pattern of branching, in contrast, vascularization is retarded (discordant maturation retardation). There is a tendency to sinusoidal dilatation; however, syncytio-­ capillary membranes are much too rare (in the present image the example displays only one arrow, compare with a. Macroscopically normal-sized placenta, sectional surface is pale (H&E, 500×)

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65 Placental Morphology

of a primary disorder in the circulation of the placenta) or may represent the primary disorder, often with a hitherto unknown ­etiology. The worst consequence of a maturation disorder is the loss of the reserve capacity of a placenta with regard to oxygenation of the fetus with consecutive intrauterine phases of fetal hypoxia, in the worst case a late intrauterine fetal death.

Intermediate Villus Deficiency A given gestational age is characterized by a typical mix of the three types of villi. When looking at the placental tissue under the microscope, the number of intermediate villi seems to be reduced in “intermediate villus deficiency”. The marked jump in caliber between the inconspicuous stem villi and an impressively greatly increased number of terminal villi (. Fig. 3.14a) with small caliber and increased vascularization is striking. The color of the placental tissue visible to the naked eye, especially when viewing the sectional surface of the parenchyma, is due to the proportion of the cross sectional area of fetal blood vessels. Although circumstances of delivery (time of cord clamping) play a role, it is generally true that a placenta will appear progressively redder as pregnancy progresses. Deviating from this, a placenta e.g. of the 28th week of gestation from a pregnancy with preeclampsia will show a dark red hue—actually only typical for a term placenta -, caused by the high proportion of fetal vessel cross-sections in the total area of the placental parenchyma. The perfusion disturbance of the placenta originating from the maternal side in preeclampsia has led to infarctions and reduced growth of the placenta. Obviously, this is compensated from the fetal side by an increase in performance through maturation with premature differentiation and increased vascularization of the terminal villi. This leads to the morphological picture of a dark red placenta and “consumption” of the growth compartment of the intermediate villi.  

Chorangiosis Chorangiosis refers to an absolute increase of blood vessels in the area of the intermediate and terminal villi. In the quantitative analysis of the cross-sections of terminal villi, up to ten vessel cross-sections per terminal villus are counted in the most severely altered areas. In this maturation disorder the functional significance of this increase is unclear. Excessive fetal vascular growth does not seem to improve the gas exchange between maternal and fetal blood, presumably because the diffusion distances remain far. Chorangiosis type I with increase in capillaries and deficient formation of syncytio-­ capillary membranes is distinguished from chorangiosis type II with sinusoidally dilated vessels and numerous syncytio-capillary membranes (. Fig.  3.15b). At the very least, chorangiosis type II appears to be a feto-­placental adaptive attempt, as placentas from mothers who go through pregnancy and delivery at high altitude have a much higher incidence of chorangiosis. Late intrauterine deaths with chorangiosis as the sole finding are very rare. The term chorangioma should be distinguished. It is a circumscribed (not diffuse) change of the placenta in the sense of a hemangioma in the chorionic villi.  

Maturation Retardation From the 34th week at the latest, the small caliber terminal villus with sinusoidal blood vessels and syncytio-capillary metabolic membranes must be by far the quantitatively dominant villous type (. Fig.  3.15a). If intermediate villi with larger caliber, higher stromal content, and only rudimentarily developed syncytio-capillary membranes are quantitatively image-dominant during this time period, the term “villous maturation retardation” is used (. Fig.  3.15c). Often the maturation-retarded placenta is large with a sectional surface that is too pale for the gestational age. This is explained by the insufficiently forced vascularization of  

 

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the villi. This disorder affecting all aspects of the morphological maturation of a placenta evenly, is typical for inadequately treated diabetes mellitus, but otherwise rare. The functional consequence is the gradual reduction of the reserve capacity of the organ for oxygenation of the fetus, whereas the nutritive function is apparently not affected. The potential danger lies in the unexpected intrauterine death of a macrosomic child, which may occur in the last six weeks of pregnancy or in the period of a prolonged pregnancy.

ity of the placenta in this respect. Therefore, terminal villus deficiency has the same functional hazard potential as diabetes-­associated maturation retardation. In two out of 1000 pregnancies, terminal villus deficiency leads to late intrauterine fetal death; the etiology is unknown. If there were an easily identifiable parameter indicating this disorder (e.g., ultrasound morphology of the placenta or hypoxia-associated measure in maternal blood), earlier induced delivery could significantly reduce perinatal mortality.

Responses and Evidence from Studies of Small-for-date Microscopic examination reveals a regular Newborns and Stillbirths

Terminal Villus Deficiency

branching of the chorionic villi with a quantitative dominance of small calibers. Only a specific analysis of the villi reveals that the morphology of a mature terminal villus is rarely found. In an ideal-typical normal terminal villus, three sinusoidally trans­ formed vessels can be recognized that occupy about 30% of the stroma and form longstretched syncytio-capillary membranes (. Fig. 3.15a). In terminal villus deficiency, capillary blood vessels are found in which a tendency to sinusoidal dilation is evident but has resulted in a regular syncytio-capillary membrane on average only once per terminal villus (. Fig. 3.15d). Since the ideal-typical terminal villi occur far too rarely, the name “terminal villus deficiency” results. As far as the maturation of the placenta is determined by the picture of villous branching, this seems to be undisturbed. Typical is the contrast to the clearly retarded vascularization, from which the term “dissociated villous maturation disorder” is derived. The entire exchange of substances between fetus and mother takes place at the surface of the chorionic villi. From the 34th week of gestation, a differently differentiated surface of the terminal villi becomes important; the syncytio-capillary membranes conduce to gas exchange. An insufficient proportion of this compartment reduces the reserve capac 

 

In most cases, a pathology of the placenta can be detected in newborns born alive but dystrophic. The need for examination results from the perceived need for explanation, possibly also from the need to support a decision made (e.g. caesarean section) by placental findings. Whether a placenta was too small for a given gestational age is easily answered using the norm chart. However, the diagnosis of placental insufficiency only arises when a pathological process within the placenta is evident. Otherwise, it could be a small placenta with a small child for other reasons (e.g. a chromosomal disorder such as uniparental disomy). The retrospective decision as to whether the fetus could have been expected to endure a further intrauterine period or whether it was high time to free it from unfavorable (placental) circumstances is hardly possible on the basis of placental morphology. Should the assumption of fetal intrauterine hypoxia conditions have led to a premature delivery, the maturation disorders (chorangiosis, maturation retardation and terminal villus deficiency) in particular should be considered in addition to the possibility of an umbilical cord complication or circumscribed premature placental abruption. Of all fetuses whose placenta has maturation retardation, few die from hypoxia in utero

67 Placental Morphology

(2.3%). However, the risk of fatal hypoxia is increased 70-fold compared to morphologically normal placentas. 3.2.5

Postpartum Period

This section deals with problems that arise only in the placental period (e.g. incomplete detachment). It also deals with the placenta as the origin of neoplasia and finally it deals with statements that can be made about twins on the basis of the placenta.

Placenta Accreta or Increta When commissioned to examine a placenta, the pathologist is rarely asked to check the completeness of the placenta; presumably because this is the classic task of the midwife and obstetrician. If manual dislodgment of the placenta has been necessary, the placenta is usually obviously incomplete or so lacerated that assessment of completeness is not possible. In this situation, the question arises whether the need for manual dislodgment can be explained by implantation pathology. The pathologist is faced with the dilemma that the most informative tissue— in the case of incomplete detachment—is probably still within the uterine cavity. An attempt is then made to examine in increased quantity those basal portions of the placenta which do not apparently correspond to an opened intervillous space. In normal implantation this region shows a zone of decidua several millimeters wide. In the basal sections of a not spontaneously detached placenta, however, smooth muscle fibers are frequently found in the immediate vicinity of placental villi (placenta accreta) or chorionic villi between uterine muscle cells (placenta increta). If this finding cannot be made, this by no means excludes a placenta accreta/increta, since—at least in the case of a clearly incomplete placenta—it can be argued that the relevant finding is still in utero.

3

Neoplasia Malignant Degeneration of the Chorial Tissue The chorionic epithelium can transform malignantly; choriocarcinoma develop in 1:25,000 to 1:40,000 pregnancies, with the relatively highest incidence observed in preceding complete hydatidiform moles. Choriocarcinoma arising from placental ­tissues grow very rapidly. However, it is easily treatable, probably because it represents a partial allograft for the maternal organism due to the paternal genetic material it contains and is therefore immunologically vulnerable. Rarely, choriocarcinoma are diagnosed in an otherwise inconspicuous placenta after term delivery. The indication for the examination of an affected placenta arises retrospectively from a focal finding which, after possible imaging as well as macroscopically, appears to be a solitary infarct. According to histological criteria, the highly atypical and highly proliferative tissue is systematically best described as an in situ choriocarcinoma. Towards the intervillous space it shows necrosis covered by fibrin, the fetal villous stroma is usually not infiltrated. Intrauterine metastasis to the fetus is very unlikely. The manifestation of choriocarcinoma in the maternal organism can typically occur even after a long latency, especially in the form of lung and/or brain metastases. Chemotherapy is still very effective even then. Differentially, placental site trophoblastic tumors, which arise from the extravillous trophoblast that physiologically infiltrates the maternal uterine wall during pregnancy, should be distinguished. This neoplasia shows low chemosensitivity. Therapy has a chance of success if complete surgical removal is successful.

Metastases to the Placenta With increasing maternal age, pregnancies are also more frequent (1:1000), in which a perhaps initially unknown malignant dis-

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ease of the mother is present. Metastasis to the placenta is very rare; by far the most common is the presence of cells of a malignant melanoma in the intervillous space, but carcinomas of the cervix, breast, lung and gastrointestinal tract also occur. By the time of birth, malignant disease has practically always become manifest in the mother. Metastasis to the fetus is even rarer; metastases of melanoma, lymphoma/leukemia, and adenocarcinoma of the lung have been observed. An indication for the examination of the placenta in case of known malignant disease of the mother can be derived from this; the metastases are usually not visible macroscopically, but have to be detected by abundant sampling. Rarely, the malignant disease may be primary in the fetus and thus cells of a neuroblastoma, a teratocarcinoma or a leukemia may be visible in the fetal circulation in the placenta. a

Twins Chorionicity and Zygosity The partition between the amniotic cavities of twins may be transparent or opaque. In the case of a common chorionic cavity (“monochorial”), the partition is transparent because it consists only of two layers of amnion (without intervening chorionic tissue) (. Fig.  3.16a). In this situation, one can read a newspaper through the stretched partition. This test for transparency proves the monochorial state and thus the monozygotism of the twins. If the fetal membranes are opaque and the newspaper cannot be read through the stretched partition, this is due to the additional chorionic tissue between the amniotic covering layers (. Fig.  3.16b). The finding is “dichorial twin pregnancy”. In this case, no statement on the zygosity is possible for the individual case.  

 

b

Fetus a

Fetus a

Fetus b Fetus b ..      Fig. 3.16  a “Monochorial means monozygotic”: If the partition between fetus A and B does not contain any chorionic tissue (dashed line), the twins originate from one zygote (“identical”). The tissue of the separating wall is slidable on the chorionic plate of the placenta and is relatively clearly transparent when stretched. b “Dichorial is not informative” (concern-

ing zygosity in individual cases): If the separating wall between fetus A and B contains chorionic tissue (dashed line), the twins may be identical or fraternal twins. The tissue of the partition is fixed on the chorionic plate of the placenta and is opaque even when stretched. (H&E, 50×)

69 Placental Morphology

Fetofetal Transfusion The dividing wall between the amniotic cavities of identical twins does not necessarily originate where the two placentas meet. Rather, the border area is a kind of watershed between the branches of the blood vessels located in the chorionic plate, which cover the chorionic plate arising from the two umbilical cord insertions. Arteries and veins may be distinguished by the fact that at the crossing points the arteries pass over the veins. Vascular connections virtually always exist between the fused placentas of identical twins. Large caliber arterio-arterial and veno-venous connections within the chorionic plate can be identified as such to the naked eye. Functionally, they can be beneficial if simultaneously existing arterio-­ venous connections in the parenchyma of the placenta form a unidirectional shunt and the resulting volume shift can be balanced out via the large-caliber connections. However, if a rapid drop in blood pressure occurs in one of the fetuses (e.g. in the case of hypoxia or premature abruption), the large-caliber connections can lead to a volume shift that is fatal for both partners in the short term. With a little practice, unidirectional shunts at the level of the villous parenchyma can be recognized by following the extensions of the vascular system in the chorionic plate. Shunts are found in areas which are supplied from one side by an arterial vessel and where a venous vessel of approximately the same size is not directed towards the same side but to the opposite side. Since in these shunts the blood can only flow from the arterial to the venous side, there will be several such areas with reversed directions of flow. Since the balance of the blood flows can hardly be the same, the large-caliber connections allow the (saving) volume balance when blood pressure differences arise. If the large connections are missing, the basis for a chronic fetofetal transfusion syndrome is laid.

3.3

3

Biobanking

Berthold Huppertz

3.3.1

Introduction

Since the beginning of the 1980s, there has been a steady increase in scientific publications on the subject of “biomarkers and the placenta”. Meanwhile, there are almost 6000 publications on the topic. This means that a large number of data and hypotheses are available on which biomarker is the better one, for which pathology it is specific and at which time it is ideally used. However, at the same time it has not been questioned, or only rarely, whether the samples used for the identification, testing and validation of these markers meet appropriate quality standards. It was not until the beginning of the 2000s that the value of systematic sample collections was recognized—and with it the necessity of a field that deals with “pre-­ analysis”. Accordingly, the increase of these publications started only after the year 2000 and has currently by far not reached the numbers of placental biomarkers (maximum 143 compared to maximum 393 for “Biomarkers and Placenta”). If only the topic “biomarkers” is taken into account, almost 60,000 publications (exactly 59,991) can be found in PubMed in 2015 alone. All of these studies should also necessarily deal with pre-­analysis.

What Exactly Is Pre-analysis? The pre-analytical phase is the period between the sample collection (e.g. blood drawing) and the analysis of the sample. The handling of the sample between these two events significantly determines the quality of the analysis result. More and more attention is being paid to quality in analyses to achieve the best possible results. Also, the era of omics technologies has made it pos-

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sible to analyze smaller and smaller amounts and more components of a sample. But do the samples used for this also meet the high demands of these technologies? Example: Imagine a serum sample in which a specific protein hormone is to be determined. This sample was taken on a morning, then treated according to the specifications in the clinic and then left in the department to be picked up by the transport service. This service overlooked the sample and did not pick it up and took it to the lab only the next day. There, the hormone level was determined and appropriate therapy was initiated for the patient. It is rather unlikely that the therapy will be adequate for the patient. This example is taken from clinical laboratory medicine, but of course applies just as much to any scientific analysis. Here, very few laboratories have a quality control system in place to check whether the samples used meet appropriate quality criteria that are necessary for the study. This is one of the reasons why in the scientific literature the data on many biomarkers are so very different and contradictory.

3.3.2

Variables Affecting the Composition of a Sample

Although the pre-analytical phase begins with the collection of the sample, other factors that influence the composition of the sample must be taken into account. Looking at the time course in . Fig. 3.17, it can be seen that even before a sample is taken (tissue, fluid, cells, etc.) a wide variety of factors can have a significant influence on the sample. Therefore, these variables are divided into two groups, the variables before and after sample collection.  

Variables Before Sampling The variables that can have a decisive influence on the sample before collection are, on the one hand, to be sought in the donor’s lifestyle and concern diet, consumption of drugs (nicotine, alcohol, etc.), intake of medication, etc.. Other variables include environmental factors such as air pollution, contact with chemicals, etc. In addition, there are variables that are directly related to the collection of samples. This is particularly true in the case of surgeries.

Time point zero The sample lives and is biologically (re)active.

Patient, Donor

Medical intervention

Removal of the Sample

The molecular composition of the sample is subject to further changes/degradations.

Further treatment/ Processing of the sample

Storage

Shipping

Scientific Analysis

Knowledge propagation

Pre-analysis Before sampling

After sampling

..      Fig. 3.17  The pre-analytical phase. Schematic representation of the pre-analytical phase between sample collection and analysis. After a sample has been

taken, a large number of variables can influence the quality and degree of preservation of a sample

71 Placental Morphology

Example: A tumor is to be removed during a surgery. For this purpose, the blood supply to the corresponding organ must be significantly restricted or stopped completely before the tissue is removed. From now on, this organ is undersupplied and runs into ischemia. Since this undersupply of the organ still occurs in the body and thus at body temperature, this is referred to as “warm ischemia”. In addition, the patient has been administered medication, is under anesthesia and the blood pressure could also vary significantly. All of these variables prior to retrieval (warm ischemia time, medication, anesthesia type and duration, etc.) affect the tumor tissue that is removed during surgery. The RNA and protein profiles of the tissue may undergo significant changes during this time due to oxygen deprivation alone, such that even immediately after removal, the tissue no longer fully resembles the tissue that was present in vivo prior to surgery. Since these variables are purely present to help the patient prior to retrieval, no scientific study should provide guidelines here. However, the relevant variables can be documented, especially the warm ischemia time. Thus, comparison of these times may explain divergent analysis data.

Variables After Sampling The pre-analytical variables after sample collection are variables that have a decisive influence on the results of all further examinations and analyses and can be considered independent of the patient. In addition, the researchers can have a direct influence on these variables. The time between sample collection and analysis is referred to as the “cold ischemia time”, as this is when the sample has left the body and is usually processed, transported and stored at ­temperatures lower than body temperature. This time, it is about sample handling, which includes the following: 55 Direct processing of the sample after collection,

3

55 Transport of the sample from the collection site to storage (or directly to analysis), 55 Storage of the sample (short, medium, long term), 55 Transport from storage to analysis, 55 Handling of the sample before analysis. During the time window between collection and analysis, a sample can be exposed to a variety of influences. This can lead to massive changes in the sample and thus significantly influence the analysis results. Since these variables are within the direct sphere of influence of the researcher, it is necessary to develop criteria before the start of sample collection that enable the best possible preservation of sample quality during the pre-­ analytical phase. If it is already known before sample collection which analyses are to be carried out, the criteria for sample handling can be specifically defined. 3.3.3

Collection or Biobank?

Researchers in the medical field have been collecting human specimens of interest for centuries. The importance of human tissue collections was politically recognized in Austria more than 200 years ago. In 1811, the Austrian Study Court Commission (Studienhofkommission) issued a decree for the “establishment and maintenance of anatomical-­ pathological cabinets”. These cabinets, i.e. anatomical-pathological collections, were to be established to promote the teaching of medical students at all medical-­ surgical teaching institutions. In addition, physicians were expressly obliged “… in all cases in which, in their clinics or in the hospitals and maternity homes entrusted to them, the opportunity arises to obtain curious anatomical-pathological specimens, games of nature, etc., to collect these or through their assistants and to hand them over to the cabinets”. The specimens in these collections were mainly fixed and preserved

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..      Table 3.1  Comparison between individual collections and biobanks

3

Individual Collection

Biobank

Collection

By an individual or a research group

Through several research groups or centrally at a clinic/university

Sample collection

For a particular study

For many possible studies not yet defined at the time of collection

Ethical vote and donor consent form

Specific to a study (if available)

Broad for use of samples/data for e.g. biomedical research

Sample diversity

Low

High

Labeling of samples

Handwritten

Barcodes

Creation of databases

Handwritten

Automated

Quality monitoring of samples and sample storage

Rarely present

Generally available

Collection of samples and documentation of data

In the hands of non-­ specifically trained persons

In the hands of appropriately trained persons

Systematic guidelines (SOPs) for sample and data handling

Rarely present

Generally available

SOP Standard Operating Procedure

with alcohol or formalin. Since—as described above—mainly “games of nature” were collected here, mainly curiosity collections developed, which can still be admired today in many anatomical collections. Towards the end of the last century, the targeted collection of further samples began, not only to fix them in formalin but also to store them frozen. Thus, the spectrum of collections expanded to include liquid samples, especially blood samples, and tissue samples stored at ultra-low temperatures (liquid nitrogen). However, it is still the individual researcher who starts these collections and supervises the storage of the samples. These collections are still operated in this way today. It was not until the 21st century that collections were taken to the next level and the term “biobank” was coined. Biobanks are fundamentally different from individual collections of researchers, as they involve a highly organized, systematic collection of

samples with their associated data that are made available for research purposes. Although biobanks have a very wide range of foci, there are fundamental differences between individual collections and biobanks. Individual collections by researchers or small groups of researchers have a number of disadvantages compared with biobanks, in which samples are collected, stored and made available for research systematically and on the basis of standardized processes (. Table 3.1).  

zz Reasons for Creating a Biobank The reasons for the emergence of biobanks arise from the following problems with individual collections. 55 Many samples that are available in individual collections cannot be used for other studies after the respective studies have been completed, because

73 Placental Morphology

55

55

55

55

no further informed consent has been obtained from the donors. The labelling of the samples is often no longer comprehensible, so that samples can no longer be assigned. Due to the manual labeling of the samples and also manual input of data, the assignment of samples to data is extremely error-prone. Data security is rarely given, since the data assigned to the samples are often stored on storage media that are not backed up multiple times. The variability of sample quality is very high: –– due to the lack of specific protocols, –– due to the lack of monitoring of sample storage, –– due to improper handling of samples (e.g. multiple freeze-thawing of blood samples).

For these and other reasons, biobanks were established in many countries to address and prevent fundamental problems in sample and data collection and storage. The first publication in PubMed on the topic of biobanking was published 1996 and by the year 2000 there were a total of 5 publications with the keyword biobank.

Definition of Biobanks To date, the term biobank is neither protected nor clearly defined. Many institutions have attempted to define it, so that today a large number of definitions can be found, some of which overlap but differ in detail. Two examples are intended to demonstrate this diversity: 55 Sweden, 2003: The Swedish Ministry of Health defined the term biobank in 2003 as “biological material from one or more human beings that is collected and preserved for an indefinite or limited period,

3

and whose origin is traceable to an individual or individuals”. 55 USA, 2016: The NIH published its ­definition of a biobank in July 2016: “A biobank is a repository that stores and manages biological samples known as biospecimens for use in research.” These two definitions illustrate the differences and overlaps very well. There is general agreement that biological material is collected and stored in biobanks—even if the species of origin of the samples is viewed differently. Biobanks generally contain biological samples. This means—in contrast to the definition in Sweden, where only human samples can be found in a biobank—that these samples can originate from plants, animals (including humans), fungi but also microorganisms. In the definition from Sweden it is not clear why these samples are collected and stored, research is missing in this definition. In addition, both definitions lack a statement about the data that can be assigned to the samples. Without these data, a sample has little or no value for research. To obtain a comprehensive definition of biobanks, the following definition can be used. >>Biobanks are facilities for the systematic collection, storage and distribution of high quality biological samples and their associated (clinical) data for research, embedded in an ethical and legal framework.

When considering human biobanks for medical research (such as clinical biobanks), these must be distinguished from collections of samples and data for other purposes. Such collections include collections for therapeutic or forensic purposes, pure medical archives for documentation, and the historical anatomical/pathological collections and museums.

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Advantages of Biobanks

3

part of this project, not only were samples from individual biobanks examined for effects during the pre-analytical phase, but interlaboratory tests were also carried out with several biobanks to identify comparabilities between biobanks. Today, the difference between individual collections and biobanks with regard to the quality of samples and data as well as ethical and legal issues has become so clear that some countries, such as Switzerland, have passed corresponding biobank laws that only permit the collection of samples and their distribution for research in and from registered biobanks. As a result of this development, biobanks are becoming increasingly important in the field of precision medicine. This significance of biobanks in the cycle of translational research is illustrated in . Fig. 3.18. It was only through the development of biobanks that the potential of omics technologies could be exploited. The data depth of these technologies revealed how crucial the pre-analytical phase is for

It was only through the development of biobanks and the implementation of corresponding quality standards that the field of new omics technologies was able to develop so rapidly. In the case of individual collections, it became apparent that the differences in quality of the samples within a collection, and especially between two or more collections, are so serious that it is no longer possible to make a clear statement about the analysis data. Analyses with samples from several collections showed that, based on the analysis data, it is easier to distinguish the collections from each other than the control samples from the disease samples. The ongoing development of quality standards in the field of biobanks is coupled with an increasing number of studies that focus on the effects of the pre-analytical phase on sample quality. One EU-funded project that has been instrumental in the development of quality standards is the SPIDIA project 7 (7 www.­spidia.­eu). As  

 

 

Biobanks

biological samples & data

Technologies

Omics technologies & imaging, Epidemiology, Bio-Informatics, Statistics

Investigative models

Patients as partners, Models of human diseases

Pathophysiological Processes

New Research questions

Cycle of translational research Clinical application

new therapies, diagnostics & markers

Technology transfer

..      Fig. 3.18  The cycle of translational research. Only the combination of high sample quality in biobanks and the development of new technologies such as omics technologies has led to a significant improve-

Identification of markers and targets

Validation of markers/ targets Screening & clinical trials

ment in the identification of markers and targets. This accelerates technology transfer and helps to find diagnoses and new therapies for patients more quickly

75 Placental Morphology

the analysis and thus for the results. Only through the combination of biobanks and omics technologies is it now possible to identify a large number of new markers and potential starting points for therapies. Subsequent validation and the corresponding technology transfer then enable precision medicine to be translated into benefits for patients (. Fig. 3.18).  

Further Readings Further Reading on Section 3.2 Becker V et al (eds) (1986) Die Plazenta des Menschen. Georg Thieme, Stuttgart

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Benirschke K, Driscoll (2011) The pathology of the human placenta. Springer, Berlin Fox H, Sebire N (2007) Pathology of the placenta. Elsevier Heerema-McKenney A (2014) Diagnostic pathology: placenta. Elsevier Stallmach T (2012) Schwangerschaft, Perinatalperiode und Kindesalter. In: Böcker et  al (Hrsg) Pathologie, 5. Aufl. Urban & Fischer & Elsevier, München, S 787–810 Stallmach T, Hebisch G (2004) Placental pathology: its impact on explaining prenatal and perinatal death (review). Virchows Arch 445:9–16 Stallmach T et al (2001) Rescue by birth: defective placental maturation and late fetal mortality. Obstet Gynecol 97(4):505–509 Vogel M (2011) Atlas der morphologischen Plazentadiagnostik. Springer, Berlin

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Placental Function— Nutrient Transport—Gas Exchange Michael Gruber, Birgit Hirschmugl, Carolin Schliefsteiner and Christian Wadsack Contents 4.1

General Functions of the Placenta – 78

4.2

Nutrient Transport Across the Placenta – 79

4.2.1 4.2.2 4.2.3 4.2.4

T ransport of Lipids and Fatty Acids – 79 Transport of Glucose – 80 Transport of Proteins and Amino Acids – 81 Transport of Minerals and Trace Elements – 83

4.3

Maternofetal Gas Exchange – 85 References – 87

© Springer-Verlag GmbH Germany, part of Springer Nature 2023 B. Huppertz, E. Schleußner (eds.), The Placenta, https://doi.org/10.1007/978-3-662-66256-4_4

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4.1

4

M. Gruber et al.

General Functions of the Placenta

Just as the placenta develops and grows during pregnancy, its functions also change during gestation. In the first three months of pregnancy, the interaction between the early placenta and the maternal decidua, which arises from the endometrium, is particularly crucial for the implantation of the embryo and for maternal tolerance to the embryo and fetus. Both the decidua and placenta contain immune cells that provide immune system adaptation to pregnancy, such as natural killer (NK) cells, macrophages, and regulatory T cells. With regard to the immunological function of the placenta, reference is made to 7 Chap. 2. The placenta is a multicellular organ and, in addition to its immunological function, also fulfils an endocrine function in that it responds to hormonal signals from the mother and child and also stimulates the production of certain pregnancy hormones which are essential for the maintenance of the pregnancy, the development of the fetus and the (timely) induction of birth. These functional aspects are discussed in 7 Chap. 5. However, the most essential functions of the placenta are the supply of nutrients to the fetus and the exchange of gases  

 

between maternal and fetal blood. These functions will be discussed in detail in the following sections. Crucial to the transport of nutrients and oxygen is the hemochorial villous architecture of the placenta, which maximizes the surface area available for exchange (7 Chap. 1), and a switch in maternal metabolism. During the first half of pregnancy, the maternal metabolism corresponds to an anabolic situation, i.e., in addition to the energy expenditure for the growth of the placenta and fetus, energy from food is primarily used to build up energy reserves (glycogen stores in the liver, fat depots). From the 20th week of pregnancy onwards, logarithmic growth of the fetus sets in and the majority of nutrients need to be made available to the child—on the one hand through increased food intake, and on the other hand through a change in maternal metabolism to a catabolic situation. Although up to three times more insulin is produced by the mother compared to the non-pregnant state (Freinkel 1980), sensitivity towards insulin decreases by about 50% (Catalano et  al. 1991; Buchanan et  al. 1990). Thereby, postprandial glucose and amino acids are supplied directly to the fetus and, in addition, energy is mobilized from maternal fat stores in the fasting state. The functions of the placenta during pregnancy are summarized in . Fig. 4.1.  

 

79 Placental Function—Nutrient Transport—Gas Exchange

until week 12

Week 13–26

Week 27 to 40

Implantation Perfusion with maternal blood First fetal blood cells and endothelial cells → start of vessel formation maternal tolerance, immunological adaptation

4

lism bo a t Ca

-

us fet e th of th w o gr id rap

Transport of glucose, amino acids, fatty acids Transport of trace elements Gas exchange

sm oli ab n A

ta en lac p the of nt e m lop ve e -d

..      Fig. 4.1  Functions of the placenta during pregnancy

4.2

 utrient Transport Across the N Placenta

4.2.1

 ransport of Lipids and Fatty T Acids

One of the main tasks of the placenta is the transport of nutrients from the maternal blood to the fetus to ensure its development and growth over the duration of pregnancy. In particular, the transport of lipids across the placenta plays an important role. In the first half of pregnancy, maternal hyperphagia leads to the build-up of maternal fat reserves in the white adipose tissue. In the course of a healthy pregnancy, the maternal lipid profile changes. Especially in the last trimester of pregnancy, increased fatty acids are mobilized from maternal lipid stores, leading to pregnancy-induced hyper-

lipidemia. This excess supply of maternal lipids is available to the fetus for growth and for the formation of its own fat reserves after transport across the placenta (Herrera et  al. 2006). Specifically, fatty acids are the major basic building blocks for the establishment of fetal lipid reserves. The transfer of fatty acids across the placenta largely follows a maternofetal concentration gradient, and only non-esterified, so-called free fatty acids are transported. However, free fatty acids account for only about 1% of the total fatty acids present in maternal plasma. The majority of fatty acids present in plasma are esterified in the form of triglycerides, phospholipids and cholesterol esters, and 99% of these are bound in lipoproteins. These fatty acids bound in lipoproteins must first be prepared for uptake into the placenta, since phospholipids and triglycerides cannot cross the placenta directly. At

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least two processes are at issue for the provision of fatty acids at the placental barrier. First, the uptake of triglyceride-rich lipoprotein particles via lipoprotein receptors on the syncytiotrophoblast and the further hydrolysis of fatty acids by intracellular lipases (Desoye et  al. 2011). Second, the release of fatty acids at the microvillous membrane of the syncytiotrophoblast by extracellular lipases, such as endothelial lipase (Gauster et  al. 2007) or lipoprotein lipase (Waterman et  al. 2000), and the uptake of the free fatty acids into the syncytium. Both diffusion and protein-mediated processes are thought to be responsible for the uptake of free fatty acids into the syncytiotrophoblast layer. In particular, the so-­ called fatty acid transport proteins (FATP 1-6, “fatty acid transport proteins”), fatty acid translocase (CD36, also FAT, “fatty acid translocase”) and intracellular fatty acid binding proteins (FABP 1-12, “fatty acid binding proteins”) are involved in the cellular uptake of long-chain free fatty acids (≥16 carbon atoms in the fatty acid chain). Whether there is a preferential interaction between these transport proteins and fatty acids with different chain length or degree of saturation is still under debate. In addition, the exact sequence of the uptake processes of fatty acids across cellular membranes has not yet been fully elucidated. However, once fatty acids from maternal plasma have entered the syncytiotrophoblast, an intermediate esterification of these fatty acids to triglycerides and phospholipids may occur. On the one hand, the phospholipids and triglycerides generated in this way represent a placental metabolic reserve, for example, for energy production by β-oxidation or for the provision of phospholipids for the expansion of cellular membranes. On the other hand, fatty acids can be hydrolyzed by lipases from placental triglycerides and phospholipids and thus be made available again for transport to the fetus (Herrera and Desoye

2016). Maternofetal transport of fatty acids favors essential fatty acids of the omega-3 and omega-6 family and their long-chain polyunsaturated fatty acid derivatives, such as docosahexaenoic acid (Haggarty 2010). This preferential transport results in an accumulation of docosahexaenoic acid in the fetus compared to maternal plasma. This observation highlights the important role of omega-3 and omega-6 fatty acids, such as docosahexaenoic acid, in fetal optic nerve and brain development (Innis 2005). However, the mechanisms underlying maternofetal fatty acid transport, including the protein factors involved, and the regulation of fatty acid transport in the placenta remain not fully elucidated and are currently the subject of numerous research projects. 4.2.2

Transport of Glucose

Glucose is an essential energy source for the basic needs of the fetus because fetal glucose production is minimal (Kalhan et al. 1979). Accordingly, the glucose concentration in maternal blood is usually higher than in fetal blood, so that the glucose flow follows a gradient from mother to child (Aynsley-­ Green et al. 1985; Bozzetti et al. 1988), with a linear relationship between the glucose concentration in maternal and fetal blood (Whaley et al. 1966; Tobin et al. 1969). The transport of glucose across the placenta is ensured by facilitated diffusion, so that no active transport that consumes additional energy is required. Nevertheless, facilitated diffusion requires glucose transporters (GLUT) that transport glucose across the microvillous and basal membranes of the syncytiotrophoblast in a Na2+ -independent manner. Three such transporters are known to exist in the placenta: GLUT1, GLUT3, and GLUT4. GLUT1 is the transporter found on all different cell types of the placenta (Hahn et  al. 1995). GLUT3 is

81 Placental Function—Nutrient Transport—Gas Exchange

expressed by fetal endothelial cells and placental stromal cells (Hahn et  al. 2001). GLUT4 is found exclusively in placental stromal cells (Xing et al. 1998). Essentially, glucose transport across the placenta is unregulated and therefore unlimited. The GLUT transport system has a high capacity and is virtually unable to saturate, which would require a glucose concentration in maternal blood of >20 mmol/l (Kalhan and Parimi 2000). Although GLUT1 is normally regulated by ambient glucose concentration, GLUT1 expression in trophoblasts in  vitro is reduced only at concentrations >20–25 mmol/l (Hahn et al. 2000). Accordingly, transport is hardly regulated by transporter availability, i.e. not by diffusion. If anything, glucose transfer to the fetus is flow-limited, which has also been shown in studies with varying flow rates of maternal and fetal blood (Gilbert et  al. 1984). It is therefore not surprising that in pregnancies with gestational diabetes, but also with fetal growth restriction, glucose transport is unchanged (Osmond et al. 2001; Challis et  al. 2000), although the level of transport molecules changes. Ex vivo perfusion of the placenta (7 Chap. 15) can be used to study the transport of various nutrients, including glucose. For placentas from diabetic pregnancies, this method has shown that glucose transport is unchanged despite the disease (Osmond et  al. 2001). Furthermore, there is also no glucose gradient between vein and arteries in the umbilical cord (Challis et al. 2000), also indicating unaltered transport. Rather, the fetal pancreas compensates for the high glucose level by overproducing insulin. However, since placental glucose transport is insulin-independent, this does not directly result in the transport of less glucose, but only in metabolizing the excess glucose, which results in a long-­term change in fetal body composition, namely the buildup of additional fat mass (Durnwald et al. 2004). Glucose transport has also been studied in  

4

fetal growth restriction (FGR, “fetal growth restriction”) using ex vivo perfusion (Challis et al. 2000). Although the placental turnover of glucose is increased in this case, the net transport is unchanged, since not only the weight of the fetus, but also the size and weight of the placenta are reduced in FGR, so that the transport remains proportionally the same (Jansson et al. 1993). Not all of the glucose transported across the placenta is consumed by the fetus per se—the placenta itself is also a metabolically very active organ and about half of the transported glucose is metabolized in the placenta and stored as glycogen or lactate (Burd et  al. 1975). The other half is thus directly available to the fetus. Although no direct measurements are possible in humans, it can be deduced from animal studies (e.g. in sheep) that the fetal rate for glucose consumption is about 5  mg glucose/kg body weight per minute (Kalhan et al. 1979). For oxidative metabolism, the glucose/O2 quotient is an essential measure to estimate whether glucose alone is sufficient to meet energy requirements. The quotient is calculated by dividing the difference in glucose levels in venous and arterial cord blood by the difference in oxygen levels in these vessels. In humans it is 0.8 (Morriss et  al. 1974)—thus glucose alone is not sufficient to meet the energy requirements of the fetus, and other sources of energy such as lactate and amino acids must be used in addition. 4.2.3

Transport of Proteins and Amino Acids

In addition to fatty acids and glucose, amino acids are an important source of energy for the growing fetus. Amino acids are not only important for building muscle mass, but are also building blocks for various biosynthetic pathways, e.g. the assembly of nucleotides and heme molecules. Impaired amino acid transport across the placenta has been asso-

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ciated in studies with reduced fetal growth (Glazier et  al. 1997; Jansson et  al. 1998), defective neurological development (Leitner et  al. 2007), and a higher risk of chronic hypertensive disease later in life (Barker 1998). The transport of amino acids across the placenta is comparatively poorly understood, but data are available from molecular biological studies (expression of transporter molecules, etc.), as well as direct data on transport rates from studies using ex  vivo perfusion of the human placenta. In contrast to glucose, the transport of amino acids must occur via “real” transporter molecules with energy expenditure. Three steps are required for this: (1) uptake of amino acids into the trophoblast via transporters at the microvillous membrane; (2) transport/turnover in the cytosol; and (3) release of amino acids at the basal membrane of the trophoblast and diffusion into the fetal circulation. Several amino acid transporters are expressed in the placenta at least at the mRNA level. Whether these mRNAs are actually transcribed to protein is unclear in many cases. Moreover, not only the presence per se, but also the subcellular localization, e.g. on the microvillous or the basal membrane of the syncytiotrophoblast, is essential. There are two main types of amino acid transporters, both of which are found in the human placenta: (1) accumulative transporters, which transport amino acids into the cell and increase the amino acid concentration in the cell, and (2) amino acid exchangers, which transport a particular amino acid only in exchange for another amino acid, thus keeping the amino acid concentration in the cell the same but changing the amino acid composition in the cell, this is essential e.g. for the uptake of essential amino acids into the cell. At the microvillous membrane of the syncytiotrophoblast, there are predominantly accumulative transporters that

actively transport amino acids against a concentration gradient into the trophoblast. These amino acids can then be further transported by amino acid exchangers in return for other amino acids. Accumulative transporters at the microvillous membrane include members of several transporter families, e.g. the system A family, which transports small neutral amino acids, or the system Y-AG, which mediates the uptake of glutamate and aspartic acid. Although the principle of uptake at the microvillous membrane is well documented, much less is known about transport across the basal membrane to the fetus. Both accumulative transporters and exchangers are present on the basal membrane. However, accumulative transporters can only take up amino acids and are therefore unlikely to be involved in the release of amino acids into the fetal circulation. Exchangers are able to mediate both uptake and release of amino acids, but can only change the composition of the amino acid pool in the cell, not the absolute concentration of amino acids. Continuous transport of amino acids to the fetus is necessary for its growth, so transporters other than amino acid exchangers that can release amino acids must theoretically be present at the basal membrane. Ex vivo placental perfusion can also be used to study the transport of amino acids. Radioactively labelled amino acids are used for this purpose. Since amino acid exchangers can only work if amino acids are present in the fetal circulation, transport across the basal membrane can be studied ­independently of exchange mechanisms by not offering amino acids on the fetal side during perfusion. After a certain time, amino acids can then be added to the fetal reservoir to additionally study transport via exchangers (Cleal et al. 2007). Such studies have demonstrated that predominantly exchangers of the alanine-serine-cysteine (ASC) and L systems are localized to the basal membrane (Kudo and Boyd 2001;

83 Placental Function—Nutrient Transport—Gas Exchange

Cleal et  al. 2007; Okamoto et  al. 2002). It has also been demonstrated that the two neutral amino acids L-serine and L-glycine, as well as L-leucine, are transported across the basal membrane independent of amino acid exchangers. Possible proteins that move amino acids across the basal membrane independent of amino acid exchange mechanisms could belong to system-N.  The recently discovered amino acid transporters TAT1, LAT3, and LAT4 could also perform this role. TAT1 and LAT4 are expressed at the mRNA level in the placenta; however, data on protein expression and exact localization are not yet available. Impaired amino acid transport has been associated with fetal growth restriction (FGR) in several studies (Mahendran et al. 1993; Jansson et al. 2002b). This particularly concerns the transport of essential amino acids (Bajoria et al. 2002; Cetin et al. 2005) such as leucine and phenylalanine (Paolini et  al. 2001). What regulatory mechanisms underlie amino acid transport that may be relevant in FGR is poorly understood. Endocrine signals from both the mother and the fetus could have a regulatory effect on transport. Furthermore, the local activity of the renin-angiotensin system appears to play a role in the regulation of blood flow to the placenta. Interestingly, changes in the concentration of angiotensin II and oxygen modulate the activity of system A-amino acid transporters in the placenta (Shibata et  al. 2006; Nelson et  al. 2003). Glucocorticoids also had similar effects on system A activity in in vitro studies (Ericsson et al. 2005). Alternatively, there is also the possibility that the placenta has a sensory function and regulates transport capacity and rates accordingly. For example, in cell culture models of isolated trophoblasts and placental explants, it has been shown that insulin, insulin-like growth factor 1 (IGF-1) and leptin have an effect on the uptake of amino

4

acids into cells. In addition to growth factors, the placenta could also act as a sensor for nutrient availability and regulate transport accordingly. Data from mouse models have shown that when amino acid availability in the mother is low, transport to the embryo is also down-regulated (Jansson and Powell 2006). Conversely, when particularly abundant nutrients are available, e.g. maternal hyperglycemia, amino acid transporters in the placenta are up-regulated (Jansson et al. 2002a). At the molecular level, the signal transduction pathway of rapamycin (mTOR) appears to be essential for nutrient sensing in many mammals; it controls cell growth depending on the availability of nutrients, e.g. branched amino acids. In the placenta, mTOR regulates the transport of L-leucine, and it has been shown that the mTOR system is down-regulated in FGR pregnancies (Roos et al. 2007), so that transport is reduced. . Figure  4.2 summarizes the most important transport pathways for macronutrients across the placenta.  

4.2.4

Transport of Minerals and Trace Elements

The transport capacity for minerals and trace elements increases with the progression of pregnancy. In the case of calcium, magnesium and phosphorus, the plasma levels reached in the newborn exceed the plasma levels of the mother (Sibley and Boyd 1988; Schauberger and Pitkin 1979). A continuous increase in iron transfer across the placenta is essential for fetal development and probably reaches a transfer rate of up to 7 mg per day in late pregnancy (Finch et  al. 1983). Placental iron uptake occurs via Fe bound to transferrin (TF)3+. A strongly increased expression of transferrin receptor 1 (TFR1) can be detected in the syncytiotrophoblast, which is only reached by cells in hematopoiesis

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M. Gruber et al.

maternal circulation A

B

Glucose CH2 OH H

4

HO

H OH H

Amino acids

O H H

C

D

E

F

VLDL

LDL

HDL

TG &PL

OH

OH

O

O

OH

HO NH2

LRP-1 GLUTlj GLIH3

1

LDLR

SR-BI

LPL, EL Syncytiotrophoblast Stroma Endothelium

2

GLIH4

fetal circulation ..      Fig. 4.2  Transport pathways for macronutrients across the placenta. a Glucose transport across the placenta follows a gradient from mother to child by “facilitated diffusion”. The glucose transporters GLUT1 (green), GLUT3 (turquoise) and GLUT4 (pink) are each found ubiquitously, in the stroma or in the endothelium. b The transport of amino acids across the placenta occurs in two steps, which depend on different transporters on the apical side (1) and the basal side (2) of the syncytiotrophoblast membrane. Accumulative amino acid transporters and exchange transporters are distinguished. c, d Triglyceride-rich lipoproteins such as VLDL and LDL bind to their respective receptors in the placenta and are taken up by receptor-mediated endocytosis. The contained triglycerides are hydrolyzed by intracellular lipases and

are available to the fetal circulation for synthesis processes or as energy storage. e, f Phospholipid- and cholesterol-rich HDL binds to the receptor SR-BI, which mediates the transport of maternal cholesterol across the placenta. Lipoprotein lipase and endothelial lipase are expressed in the syncytium and may also contribute to the release of free fatty acids and phospholipids from lipoproteins. VLDL Very low density lipoprotein, LDL Low density lipoprotein, HDL High density lipoprotein, TG Triglycerides, PL Phospholipids, LPL Lipoprotein lipase, EL Endothelial lipase, LRP-1 Low density lipoprotein receptor-related protein 1, LDLR Low density lipoprotein receptor, SR-­BI Scavenger receptor class B type I. (Adapted from Desoye et al. 2011)

(Ponka and Lok 1999). The Fe3+ -TF complex is taken up into the syncytium after binding with TFR1 in endosomes. The exact mechanism of the reduction of Fe3+ to Fe2+ in the syncytium is unknown. However, it is suspected that STEAP family proteins, which are detectable in the placenta, catalyze this reduction in the placenta, as such reduction has been shown in other cell types (Ohgami et  al. 2006). The Fe2+ is excreted from the endosome by the transport protein “divalent metal transporter 1” (DTM1) or a protein of the zinc transporter family (ZIP) and transported intracellularly in the syncytium (Fleming et  al. 1998; Pinilla-Tenas

et  al. 2011). Transport from the syncytium into the placental stroma and further transport into the fetal circulation is thought to be accomplished by ferroportin (Drakesmith et  al. 2015) and ferritin (Maymon et  al. 2000). Calcium is required for the mineralization of the fetal skeleton and reaches a transport capacity of 140  mg/kg body weight per day at the end of the third trimester (Salle et  al. 1987). This transport rate results in an amount of calcium of up to 30  g being transported across the placenta by the end of pregnancy (Lafond et  al. 2001). This is achieved by sodium-calcium

85 Placental Function—Nutrient Transport—Gas Exchange

pumps (Belkacemi et al. 2005) and calcium ATPases (Moreau et al. 2003) in the apical syncytiotrophoblast membrane. A calcium concentration gradient between maternal blood (2.23  mM) and the syncytium (0.1–1 μM) also has a favorable effect on calcium flux (Husain and Mughal 1992). This gradient is thought to be maintained by calcium-­binding proteins such as calbindin-­ D-­28 k (Belkacemi et al. 2003) and chaperone proteins such as clanexin (Tjoelker et al. 1994). A simultaneous buffering or storage function of these proteins is assumed to ensure a continuous supply of calcium to the fetus. There is little evidence on how magnesium is supplied to the fetus via the placenta. It can be assumed that this is a calcium-­ independent energy-consuming process (Lourdes et al. 1992). In a mouse model, the magnesium transport protein TRPM6 has been linked to pregnancy outcome (Chubanov et  al. 2016). In humans, a link between defects in the TRPM6 protein and hypomagnesemia has been shown (Voets et al. 2004). Overexpression of the SLC41A1 gene in preeclamptic placentas has been described and may suggest a key function of this magnesium transport protein in magnesium homeostasis (Kolisek et  al. 2013). Further research in the field of placental magnesium balance will be necessary to adequately describe it. Transport of phosphorus across the placenta is strongly sodium-associated, as in vitro studies have shown. Likewise, a pH and oxygen dependence has been demonstrated in the experiments (Lajeunesse and Brunette 1988; Lafond et  al. 1988). The mRNA of two sodium-dependent phosphorus transporters, Slc20a1 and Slc20a2, has been detected in the placenta (Nishimura and Naito 2008). A detection at the protein level as well as an expression difference in preeclampsia substantiates the assumption that these two transporters are relevant in pregnancy (Yang et al. 2014).

4

The syncytiotrophoblast is thought to be the main regulator of zinc uptake (Mas and Sarkar 1991). A potassium-zinc transporter has been detected in the syncytial membrane (Aslam and McArdle 1992). Expression of mRNA for the following SLC30 family transporters has been demonstrated for the human placenta: ZnT1, 2, 4, and 5 (Ford 2004). As with iron transport, involvement of ZIP proteins in zinc transport is inferred from strong expression (Wang et  al. 2012). Likewise, uptake of zinc bound to alpha-­2-­ microglobulin into the placenta has been demonstrated (Douglas et  al. 1998). The presence of the metal binding protein metallothionein in fetal amniotic and villous interstitial cells as well as the syncytiotrophoblast suggests that this protein is involved in the transplacental transport of zinc (Goyer et al. 1992). The regulation and fetal release of zinc are the subject of current research and are not fully understood.

4.3

Maternofetal Gas Exchange

The respiratory gases oxygen and carbon dioxide can be considered the most important gases to be transported across the placenta. Gas exchange is significantly influenced by hemoglobin. The hemoglobin molecule consists of a heme group attached to the protein globin, the final molecule is a tetrameter and consists of four subunits. The main amino acid sequences of globin in human development are the five different chains, α-, β-, γ-, ε- and ζ-. The combination of these chains yields the differences between embryonic hemoglobin Gower-1 ζ2 ε2, Gower-2 α2 ε2, Portland-­1 ζ2 γ2, Portland-2 ζ β22, fetal hemoglobin (HbF) α γ22, and adult hemoglobin (HbA) α2 β2. All hemoglobins can adopt two conformations. The deoxygenated or t-form (“tense”) and the oxygenated or r-form (“relaxed”). Oxygenation of one heme

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group facilitates oxygenation of the remaining heme groups and results in a conformational change from t- to r-form. Crystallography shows that hemoglobin is more compact in the r-form (Perutz 1964). The two conformations can explain interactions with 2,3-bisphosphoglycerate (Perutz 1970) and the Bohr effect, which affect the oxygen binding affinity. The hemoglobin-oxygen interaction depends on the oxygen saturation of the hemoglobin and the oxygen partial pressure. If these two terms are plotted on a graph, the sigmoidal equilibrium curve of oxygen is obtained, which is summarized by the term P50. P50 describes a 50% saturation of hemoglobin with oxygen at pH 7.4 and body temperature. This P50 saturation is already reached at an oxygen partial pressure of 27 mmHg with HbA (Delivoria-Papadopoulos and McGowan 2011). The sigmoidal shape of the oxygen saturation curve of hemoglobin can only be obtained if both of the following assumptions are made: 55 the heme groups interact with oxygen in a defined sequence, and 55 the heme groups influence each other in oxygen binding. A regularity for this oxygen saturation property of hemoglobin was first described by Hill in the following equation: y / 100 =

Kx n 1 + Kx n

In the equation, y refers to the oxygen saturation in %, K describes the equilibrium constant, x corresponds to the oxygen partial pressure and n to the average number of iron atoms involved in oxygen binding (Hill 1910). Normal HbA has an n value of approximately 2.9. HbA has a theoretical oxygen binding capacity of 1.39 ml of oxygen per gram of hemoglobin, which is not reached physiologically. The real oxygen binding capacity of hemoglobin was deter-

mined by the German chemist Gustav Hüfner to be 1.34 ml of oxygen per gram of hemoglobin under normal conditions (von Hüfner 1889). Another important effect for gas exchange is the Bohr effect. This describes the dependence of the oxygen binding capacity of hemoglobin in relation to the pH value or CO2 partial pressure. Bohr et al. were able to show that at higher CO2 partial pressures, hemoglobin binds less oxygen than at low CO2 partial pressures (Bohr et al. 1904). The lower pH favors the portioning of histidine residues in the hemoglobin structure, leading to stabilization of the t-form of hemoglobin, which explains the decreased oxygen binding affinity (Kovalevsky et  al. 2010). Organic phosphates, most notably 2,3-bisphosphoglycerate (2,3-BPG), have an effect on the oxygen affinity of hemoglobin (Benesch and Benesch 1967; Chanutin and Curnish 1967). 2,3-BPG binds to deoxyhemoglobin in a molar ratio of 1:1, thus favoring the t conformation. It has a stabilizing effect on the t-form, resulting in reduced oxygen affinity (Arnone 1972). 2,3-BPG binds differently to the different hemoglobin variants. An increase in 2,3-BPG concentration during pregnancy in fetal and maternal erythrocytes may also influence gas exchange across the placenta in favor of the fetus, since fetal hemoglobin has a lower affinity for 2,3-BPG (Delivoria-Papadopoulos et al. 1971). The differences in the chain structure of hemoglobin are reflected in its oxygen binding properties. . Table  4.1 gives an overview of the different physiologically occurring hemoglobin variants during human development. Embryonic and fetal hemoglobin has a higher oxygen binding capacity than adult hemoglobin, which can be seen in the Hill coefficient and P50 oxygen saturation. Another difference between HbF and HbA is the difference in the Bohr effect. Oxygen  

4

87 Placental Function—Nutrient Transport—Gas Exchange

.       Table 4.1  Properties of the different hemoglobin variants in vitro Hemoglobin

Hill Coefficient

P50 in vitro (mmHg)

ζ2ε2

1.7

α 2 ε2

2,3-BPG Binding Constant (mM)

Bohr Effect (Δlog P50/ ΔpH)

Reference

1.4

0.09

−0.25

He and Russell (2001)

2.3

2.7

0.17

−0.51

He and Russell (2001)

ζγ2

2.3

5.9

6.0

−0.3

Hofmann et al. (1995)

ζ2β2

1.6

1.9

0.30

−0.1

He and Russell (2001)

α2 γ2

2.65

−0.51

Zhang et al. (2003)

α2 β2

2.9

−0.54 (−0.41)

He and Russell (2001) and Hofmann et al. (1995)

2

P50 in vivo (mmHg)

20 3.2 (9.8)

27

binding is more reduced in HbA than in HbF when CO2-partial pressure increases. This also favors oxygen transport from the mother to the fetus. Compared to HbA, HbF has a low P50 oxygen saturation of 20  mmHg instead of 31 mmHg in the third trimester. This means that at a usual oxygen partial pressure of 30  mmHg in the umbilical vein, there is a 6-8% higher loading of fetal hemoglobin compared to maternal hemoglobin (Beer et al. 1958).

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Portland-2 (zeta2beta2) assembled in complex transgenic-knockout mice. Blood 97(4):1099– 1105 Herrera E, Desoye G (2016) Maternal and fetal lipid metabolism under normal and gestational diabetic conditions. Horm Mol Biol Clin Investig 26(2):109–127 Herrera E et al (2006) Maternal lipid metabolism and placental lipid transfer. Horm Res 65(S3):9–64 Hill AV (1910) The possible effects of the aggregation of the molecules of hæmoglobin on its dissociation curves. J Physiol 40(Supplement-Proceedings of the physiological society):iv–vii Hofmann O, Mould R, Brittain T (1995) Allosteric modulation of oxygen binding to the three human embryonic haemoglobins. Biochem J 306(Pt 2):367–370 Husain SM, Mughal MZ (1992) Mineral transport across the placenta. Arch Dis Child 67:874–878 Innis SM (2005) Essential fatty acid transfer and fetal development. Placenta 26(Suppl A):70–75 Jansson T, Powell TL (2006) IFPA 2005 Award in placentology lecture. Human placental transport in altered fetal growth: does the placenta function as a nutrient sensor?—a review. Placenta 27 Suppl A:91–97 Jansson T, Scholtbach V, Powell TL (1998) Placental transport of leucine and lysine is reduced in intrauterine growth restriction. Pediatr Res 44(4):532– 537 Jansson T, Wennergren M, Illsley NP (1993) Glucose transporter protein expression in human placenta throughout gestation and in intrauterine growth retardation. J Clin Endocrinol Metab 77(6):1554– 1562 Jansson T et al (2002a) Alterations in the activity of placental amino acid transporters in pregnancies complicated by diabetes. Diabetes 51(7):2214– 2219 Jansson T et al (2002b) Glucose transport and system A activity in syncytiotrophoblast microvillous and basal plasma membranes in intrauterine growth restriction. Placenta 23(5):392–399 Kalhan S, Parimi P (2000) Gluconeogenesis in the fetus and neonate. Semin Perinatol 24(2):94– 106 Kalhan SC et  al (1979) Glucose production in pregnant women at term gestation. sources of glucose for human fetus. J Clin Invest 63(3):388–394 Kolisek M et  al (2013) Hypertension in pregnancy SLC41A1 is the only magnesium responsive gene significantly overexpressed in placentas of preeclamptic women. Hypertension Pregnancy 32(4):378–389 Kovalevsky AY et  al (2010) Direct determination of protonation states of histidine residues in a 2 A neutron structure of deoxy-human normal adult

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hemoglobin and implications for the Bohr effect. J Mol Biol 398(2):276–291 Kudo Y, Boyd CAR (2001) Characterisation of L-tryptophan transporters in human placenta: a comparison of brush border and basal membrane vesicles. J Physiol 531(2):405–416 Lafond J et al (1988) Parathyroid hormone receptor in human placental syncytiotrophoblast brush border and basal plasma membranes. Endocrinology 123(6):2834–2840 Lafond J et al (2001) Hormonal regulation and implication of cell signaling in calcium transfer by placenta. Endocrine 14(3):285–294 Lajeunesse D, Brunette MG (1988) Sodium gradient-­ dependent phosphate transport in placental brush border membrane vesicles. Placenta 9(2):117–128 Leitner Y, Fattal-Valevski A, Geva R, Eshel R, Toledano-­ Alhadef H, Rotstein M, Bassane H, Radianu B, Bitchonsky O, Jaffa AJ et  al (2007) Neurodevelopmental outcome of children with intrauterine growth retardation: a longitudinal, 10-Year prospective study. J Child Neurol 22:580– 587 Lourdes M et  al (1992) Effect of chronic maternal dietary magnesium deficiency on placental calcium transport. J Am Coll Nutr 11(1):87–92 Mahendran D et  al (1993) Amino acid (system A) transporter activity in microvillous membrane vesicles from the placentas of appropriate and small for gestational age babies. Pediatr Res 34(5):661–665 Mas A, Sarkar B (1991) Binding, uptake and efflux of 65Zn by isolated human trophoblast cells. BBA— Mol Cell Res 1092(1):35–38 Maymon R et al (2000) Localization of p43 placental isoferritin in human maternal-fetal tissue interface. Am J Obstet Gynecol 182(3):670–674 Moreau R et al (2003) Expression and role of calcium-­ ATPase pump and sodium-calcium exchanger in differentiated trophoblasts from human term placenta. Mol Reprod Dev 65(3):283–288 Morriss FH et al (1974) The glucose/oxygen quotient of the term human fetus. Biol Neonate 25(1– 2):44–52 Nelson DM et  al (2003) Hypoxia reduces expression and function of system A amino acid transporters in cultured term human trophoblasts. AJP: Cell Physiol 284(2):C310–C315 Nishimura M, Naito S (2008) Tissue-specific mRNA expression profiles of human solute carrier transporter superfamilies. Drug Metab Pharmacokinet 23(1):22–44 Ohgami RS et al (2006) The steap proteins are metalloreductases. Blood 108(4):1388–1394 Okamoto Y et al (2002) Expression and regulation of 4F2hc and hLAT1 in human trophoblasts. Am J Physiol Cell Physiol 282(1):C196–C204

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Osmond DT et al (2001) Placental glucose transport and utilisation is altered at term in insulin-treated, gestational-diabetic patients. Diabetologia 44(9):1133–1139 Paolini CL et al (2001) Placental transport of leucine, phenylalanine, glycine, and proline in intrauterine growth-restricted pregnancies. J Clin Endocrinol Metab 86(11):5427–5432 Perutz MF (1964) The hemoglobin molecule. Sci Am 211:64–76 Perutz MF (1970) Stereochemistry of cooperative effects in haemoglobin. Nature 228:726–734 Pinilla-Tenas JJ et al (2011) Zip14 is a complex broad-­ scope metal-ion transporter whose functional properties support roles in the cellular uptake of zinc and nontransferrin-bound iron. AJP: Cell. Physiology 301(4):C862–C871 Ponka P, Lok CN (1999) The transferrin receptor: role in health and disease. Int J Biochem Cell Biol 31(10):1111–1137 Roos S et al (2007) Mammalian target of rapamycin in the human placenta regulates leucine transport and is down-regulated in restricted fetal growth. J Physiol 582(1):449–459 Salle B et al (1987) Vitamin D metabolism in preterm infants. Biol Neonate 52:119–130 Schauberger CW, Pitkin RM (1979) Maternal-­ perinatal calcium relationships. Obstet Gynecol 53(1):74–76 Shibata E et al (2006) Angiotensin II decreases system A amino acid transporter activity in human placental villous fragments through AT1 receptor activation. Am J Physiol Endocrinol Metab 291(5):E1009–E1016 Sibley CP, Boyd RD (1988) Control of transfer across the mature placenta. Ox Rev Reprod Biol 10:382– 435 Tjoelker LW et al (1994) Human, mouse, and rat calnexin cDNA cloning: identification of potential

calcium binding motifs and gene localization to human chromosome 5. Biochemistry 33(11):3229– 3236 Tobin JD, Roux JF, Soeldner JS (1969) Human fetal insulin response after acute maternal glucose administration during labor. Pediatrics 44(5):668– 671 Voets T et  al (2004) TRPM6 forms the Mg2+ influx channel involved in intestinal and renal Mg2+ absorption. J Biol Chem 279(1):19–25 von Hüfner G (1889) Ueber krystallinisches Hämaglobin. Zeitschrift für physiolog. Chemie, XI–XIII Wang CY et al (2012) ZIP8 is an iron and zinc transporter whose cell-surface expression is up-­ regulated by cellular iron loading. J Biol Chem 287(41):34032–34043 Waterman IJ et al (2000) Further characterization of a novel triacylglycerol hydrolase activity (pH 6.0 optimum) from microvillous membranes from human term placenta. Placenta 21(8):813–823 Whaley WH, Zuspan FP, Nelson GH (1966) Correlation between maternal and fetal plasma levels of glucose and free fatty acids. Am J Obstet Gynecol 94(3):419–421 Xing AY et  al (1998) Unexpected expression of glucose transporter 4  in villous stromal cells of human placenta. J Clin Endocrinol Metab 83(11):4097–4101 Yang H et  al (2014) Comparing the expression patterns of placental magnesium/phosphorus-­ transporting channels between healthy and preeclamptic pregnancies. Mol Reprod Dev 81(9):851–860 Zhang Y et al (2003) The cooperativity of human fetal and adult hemoglobins is optimized: a consideration based on the effectiveness of the Bohr shift. Zoolog Sci 20(1):23–28

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Endocrinology of the Placenta Ekkehard Schleußner Contents 5.1

Introduction – 92

5.2

Steroid Hormones – 96

5.2.1 5.2.2 5.2.3

 rogesterone – 96 P Estrogens – 96 Glucocorticoids – 97

5.3

Peptide Hormones – 98

5.3.1 5.3.2 5.3.3 5.3.4

 uman Chorionic Gonadotropin (hCG) – 98 H Leptin – 99 Corticotrophin Releasing Hormone (CRH) – 100 Placental Lactogen (hPL) and Placental Growth Hormone (hPGH) – 102 Summary – 103

5.3.5

References – 103

© Springer-Verlag GmbH Germany, part of Springer Nature 2023 B. Huppertz, E. Schleußner (eds.), The Placenta, https://doi.org/10.1007/978-3-662-66256-4_5

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5.1

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Introduction

The placenta is the largest endocrine organ during pregnancy dominating the entire hormonal system. Since the placenta is not innervated, all interaction with the mother as well as the fetus must be performed through hormonal signals. However, these “communication signals” secreted by the placenta into the maternal and fetal blood circulation can also act locally in the paracrine and autocrine regulation of placental function. Placental peptide and steroid hormones influence all maternal endocrine functional circuits and regulatory axes. Even the not yet implanted embryo shortly after fertilization sends hormonal signals, especially human chorionic gonadotropin (hCG), which lead to the maintenance of the progesterone-­producing corpus luteum in the ovary and its transformation into the corpus luteum graviditatis over the first ten weeks of pregnancy. At the same time, successful implantation into the uterine wall and early placentation are mediated in this way and via paracrine signals. The hormones are synthesized mainly in the syncytiotrophoblast and the villous cytotrophoblast of the placental villi, but also in the extravillous trophoblast. These

hormones can be subdivided according to their biochemical structure into (. Table 5.1): 55 Steroid hormones are synthesized or modified in the trophoblast from maternal cholesterol, but also from fetal precursor hormones (. Fig.  5.1). Their synthesis thus embodies the concept of the fetoplacentomaternal unit established almost 50 years ago (Diczfalusy 1984). As lipophilic hormones, they are readily membrane-permeable and act at intracellular and nuclear receptors. However, membrane-bound receptors have also been described. In the following, the effects of progesterone, estrogen and glucocorticoids are highlighted. 55 In addition to the placenta-specific hormones human chorionic gonadotropin (hCG), relaxin, human placental lactogen (hPL) and human placental growth hormone (hPGH), peptide hormones include a large number of other hormones and endocrine factors that are either identical to hormones occurring in endocrine homeostasis such as corticotrophin releasing hormone (CRH), leptin, gonadotropin releasing hormone (GnRH), etc. or are active at their receptors such as hCG at the gonadotropin receptors (Petraglia et al. 1996) (. Table 5.1).  

 

 

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93 Endocrinology of the Placenta

..      Table 5.1  Overview of the most important placental hormones and their function. (Modified according to Costa 2016) Hormone

Source

Receptor

Function

Cellular Regulation

Progesterone

Syncytiotrophoblast

Nuclear receptor Membrane-­ associated receptor

Decidualization Embryo implantation Tranquilizing the myometrium Developing immunotolerance through –  PIBF –  ↑ Th2 cytokine production –  ↓ uNK cell activity ↓ Trophoblast invasion ↑ Trophoblast migration ↓ Placental synthesis of hCG and leptin

PKA (↑) Estradiol (↑) Insulin (↑) IGF-1 (↑) Calcitriol (↑) Leptin (↓) CRH (↓) p38, ERK 1/2 (↑ 3βHSD)

Estradiol Estriol Estrone

Syncytiotrophoblast

Nuclear receptor Membrane-­ associated receptor

Endometrial receptivity and embryo implantation Angiogenesis and regulation of uteroplacental perfusion ↑ Myometrial contractility and induction of labor Growth of the mammary glands and preparation for lactation Syncytialization ↑ Placental leptin synthesis Hyperlipidemia and fat storage

cAMP (↑ aromatase) hCG (↑ aromatase) Estradiol (↑ aromatase) Cortisol (↑ aromatase) ERK 1/2 (↓ aromatase) CRH (↑) Leptin (↓) Insulin (↓)

hCG (human Chorionic Gonadotropin)

Syncytiotrophoblast

LH/hCG receptor

↑ Progesterone production in the corpus luteum graviditatis Syncytialization ↑ Decidual angiogenesis Support of immunotolerance ↑ Trophoblast invasion Tranquilization of the myometrium Embryo implantation

cAMP/PKA (↑) p38, ERK 1/2 (↑) PPAR-γ (↑) GnRH (↑) EGF (↑) Leptin (↑) Progesterone (↓) Activin (↑) Inhibin (↓)

hPL (human Placental Lactogen)

Syncytiotrophoblast Extravillous trophoblast

GH receptor Prolactin receptor

Lipolysis ↑ Free fatty acids and ketones ↑ Insulin sensitivity ↑ Fetal insulin and IGF-1 synthesis ↓ Placental leptin synthesis

cAMP (↑) GHRF (↑) Insulin (↑) EGF (↑) PPAR-γ (↑) Calcitriol (↑) Apolipoproteins (↑) IL-1, IL-6 (↑)

Steroid hormones

Peptide Hormones

(continued)

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..      Table 5.1 (continued)

5

Hormone

Source

Receptor

Function

Cellular Regulation

hPGH (human Placental Growth Hormone)

Syncytiotrophoblast Extravillous trophoblast

GH receptor

↑ Maternal IGF-1 synthesis ↑ Lipolysis Insulin resistance ↑ Blood glucose level ↑ Trophoblast invasion

cAMP (↑) Hypoglycemia (↑) Visfatin (↑) PPAR-γ (↑) Insulin (↓) Leptin (↓) Cortisol (↓)

Leptin

Syncytiotrophoblast Extravillous trophoblast

Leptin receptor

↑ Cytotrophoblast proliferation ↓ Cytotrophoblast apoptosis ↑ Trophoblast invasion ↑ hCG and ↓ hPGH synthesis ↓ Progesterone and estradiol production ↑ Angiogenesis Embryo implantation Uterine immunomodulation Tranquilization of the myometrium ↑ Proinflammatory cytokines and prostaglandins

cAMP/PKA (↑) PKC (↑) p38, ERK 1/2 (↑) hCG (↑) Estradiol (↑) Insulin (↑) Progesterone (↓) hPL (↓) Hypoxia (↓)

CRH (Corticotrophin Releasing Hormone)

Syncytiotrophoblast

CRH receptor 1 and 2

↑ Remodeling by matrix metalloproteinase 9 (MMP-9) ↑ Trophoblast invasion and growth Regulation of birth initiation Stimulation of prostaglandin effects by –  ↑ 15-OH-­prostaglandin dehydrogenase (PGDH) –  ↑ Prostaglandin synthase (PGHS)-2 expression –  ↓ Progesterone release Stimulation of maternal and fetal adrenergic regulation ↑ Cortisol and DHEAS concentration ↑ Placental estrogen synthesis

Neurotransmitter (NA, Ach) (↑) Neuropeptides –  Angiotensin II (↑) –  Vasopressin (↑) Interleukin-1 (↑) Glucocorticoids (↑) Progesterone (↓) NO (↓)

Adiponectin

Syncytiotrophoblast

Adiponectin receptor 1 and 2

↓ hCG, progesterone and hPL secretion ↓ Cytotrophoblast proliferation Syncytialization ↓ Placental insulin signaling ↑ Trophoblast invasion Modulation of uteroplacental angiogenesis ↑ Proinflammatory cytokines

Leptin (↓)? TNF-α (↓)? IL-6 (↓)?

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5

..      Table 5.1 (continued) Hormone

Source

Receptor

Function

Cellular Regulation

Activin A

Syncytiotrophoblast

Activin receptor IIa+b

↑ Cytotrophoblast proliferation ↑ Trophoblast invasion Decidualization and endometrial receptivity ↑ hCG and progesterone secretion ↑ Placental aromatase activity

Oxidative stress (↑) Proinflammatory cytokines (↑) CRH (↑) Hypoxia (↓)

Inhibin A

Syncytiotrophoblast

Activin receptor IIa+b

Antagonization of the Activin effects hCG (↑) cAMP (↑) EGF (↑) GnRH (↑) Prostaglandin (↑) Activin (↓) Hypoxia (↓)

PKA Protein kinase A, PKC Protein kinase C, PIBF Progesterone-induced blocking factor, IGF-1 Insulin-like growth factor 1, ERK Extracellular signal-regulating kinase, cAMP Cyclic adenosine monophosphate, PPAR-y Peroxisome proliferator activating receptor gamma, GnRH Gonadotropin releasing hormone, EGF Epidermal growth factor, GHRF Growth hormone releasing factor, PHG Placental growth hormone, NO Nitric oxide, TNF-α Tumor necrosis factor alpha.

Maternal

Placenta

Fetal

Adrenal Cortex CYP450scc

CYP450scc

Pregnenolone sulfate

Pregnenolone 3ß-HSD

Adrenal gland

Progesterone Androstenedione 3ß-HSD

DHEAS

Aromatase

Liver

Estrone 16α-OH-DHEAS

17S-HSD 17ß-estradiol Estriol

se ata m o Ar

..      Fig. 5.1  Steroid synthesis in the feto-placento-­ dehydrogenase/Δ5-Δ4-isomerase, 17β-HSD 17-Beta-­ maternal unit. CYP450scc cytochrome P450 “side-­ hydroxysteroid dehydrogenase, DHEAS Dehydroepichain cleavage”; 3β-HSD 3-Beta-hydroxysteroid androsterone

96

Steroid Hormones

ways involved in proliferation, differentiation as well as angiogenesis and ultimately decidualization. Genomic progesterone effects are 5.2.1 Progesterone thus crucial for nidation and implantation of Progesterone is the central pregnancy-­ the embryo (Halasz and Szekeres-Bartho maintaining hormone. It is necessary for the 2013). In addition, progesterone also mediates non-­ genomic effects through relaxation of the myometrium, which enables rapid, membrane-­ p ositioned receptors (MPRs) that a non-soft uterus free of contractions. At the decrease cAMP production through MAPK same time it has anti-­ inflammatory and 2+ immunosuppressive functions, which enable activation and regulate intracellular Ca the formation of the necessary immune tol- release (Goldman and Shalev 2007). Progesterone is instrumental in the develerance at the fetomaternal interface. opment of fetomaternal immune tolerance Progesterone is initially formed in the by enhancing the expression of Th2 cytocorpus luteum graviditatis and, by decidualkines and inhibiting the activity of uterine izing the endometrium, creates the prerequinatural killer T cells (uNK). Their cytotoxsite for nidation and implantation of the icity is inhibited by progesterone-induced embryo. Both the maintenance of the corpus luteum graviditatis and its synthesis are blocking factor (PIBF) (Halasz and controlled by hCG secreted by the concep- Szekeres-­Bartho 2013). Inhibition of proinflammatory Th1 cytotus and are thus the result of fetomaternal kines is also a mechanism for maintaining th communication. After the 8 week of gestauterine tranquilization by decreasing prostation, a luteoplacental shift takes place and glandin and NF-κB synthesis. Progesterone the placenta takes over the synthesis of proalso inhibits myometrial contractility gesterone in sufficient quantity to maintain through the expression of proteins essential pregnancy (Tuckey 2005). for labor, such as matrix metalloproteases, Progesterone is converted in the syncy2+ channels, connexin 43 (a component of Ca tium from maternal cholesterol by cytochrome P450scc (“side-chain-cleavage”) to gap junctions relevant to labor synchrony), Δ5-pregnenolone and then to progesterone and oxytocin receptors (Kuon et  al. 2015). by means of the enzyme 3-beta-­hydroxysteroid Progesterone also acts as an a­ ntagonist of dehydrogenase/Δ5-Δ4-­isomerase (3β-­HSD) CRH-triggered induction of delivery. 5.2

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type I.  The 3β-HSD activity is the limiting synthesis step in this process (. Fig. 5.1). Maternal blood progesterone levels increase during pregnancy until the last weeks before birth, but there is no decrease before the onset of labor in humans as in other species (Csapo et al. 1971). Progesterone binds to both classical nuclear PRα and PRβ receptors, which are ubiquitously present throughout the genital tract and placenta, as well as other organs such as the mammary glands and brain. Upon binding, steroid receptors dimerize and exert their genomic effects in the nucleus by binding to specific promoter regions. In addition, PRβ can also activate cytoplasmic signaling path 

5.2.2

Estrogens

Estrogens are synthesized in the ovary, the adrenal glands and the placenta. During the first weeks of pregnancy, estradiol synthesis is maintained in the corpus luteum graviditatis under the influence of hCG, before the syncytiothrophoblast takes over the production in large quantities, which increases over the entire course of pregnancy. A group of estrogens is formed in the placenta: Estrone (E1), 17β-estradiol (E2), estriol (E3), and estrol (E4), but only E2 and E3 have clinical significance.

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However, the placenta is unable to carry tion across the uterus. This mechanism plays out the synthesis step of 17-hydroxylation an essential role in the induction of labor at of C21 steroids and therefore requires the term as well as in threatened preterm birth precursors dehydroepiandrosterone sulfate (Di et al. 2001). (DHEAS) for estradiol and 16-hydroxy-­ During pregnancy, however, estradiol DHEAS for estriol synthesis. DHEAS is uti- plays a far more important role, both syslized by both maternal and predominantly temically and locally. Since it is secreted in fetal adrenal cortex and is further aroma- large quantities into the maternal circulatized to estrogens (. Fig.  5.1). Placental tion, it plays an essential role in the adaptaCYP450 aromatase activity is oxygen-­ tion of the maternal organism to pregnancy. dependent, so that estrogen production is Estrogen-induced changes in homeostasis decreased in placental insufficiency. In con- during pregnancy lead, among others, to trast, during fetal stress, elevated levels of fluid retention and increase in blood volume, estriol are found in maternal blood because hypercoagulability, inhibition of lipolysis its sole precursor, 16-OH-DHEAS, is and hyperlipidemia, as well as to the growth derived from the activated fetal stress axis in of uterine and mammary gland tissues in such cases (Diczfalusy 1984). Before fetal preparation for lactation (Schleußner 2017). status diagnosis by cardiotocography (CTG) Estradiol regulates uteroplacental angioand sonography was established, this was genesis and endothelial proliferation via used to detect fetal compromise in antenatal ERα and ERβ receptors, as well as uterine care. Placental estrogen synthesis is lower in and placental vasodilation via nongenomic multiparous women and in pregnant women effects (Corcoran et  al. 2014). But even >30 years than in primigravidae and younger before that, estradiol supports embryo women (Toriola et al. 2011). implantation by stimulating endometrial Estrogens act firstly via their nuclear growth and differentiation, as well as cytoreceptors ERα and ERβ, which, like proges- trophoblast differentiation and syncytializaterone receptors dimerize after binding and tion via ERα (Groothuis et  al. 2007; modulate gene expression. ERα receptors Bukovsky et  al. 2003). At the end of pregare predominantly expressed in the cytotro- nancy, estradiol, like estriol, induces the forphoblast, whereas ERβ receptors are more mation of gap junctions and, via ERα expressed in the syncytiotrophoblast. On the activation, the expression of oxytocin recepother hand, membrane-bound estrogen tors in the myometrium, thereby increasing receptors exert nongenomic effects on e.g. its contractility (Renthal et al. 2015). activation of adenylate cyclase, intracellular Ca2+ mobilization and MAPK activity (Bjornstrom and Sjoberg 2005). 5.2.3 Glucocorticoids Estriol, which is derived exclusively from fetal precursors, is considered a weak estro- The placenta does not synthesize glucocortigen but can be measured well in maternal coids de novo, but regulates fetal exposure urine. It has vasodilating effects on the through maternal stress hormones. uteroplacental vasculature and can thus Glucocorticoids play a central role in fetal optimize fetoplacental supply (Chang and organ development and maturation, so that Zhang 2008). At the same time, it increases the fetus must be protected from excessive gap junction formation in the myometrium maternal levels, which may have effects on via increased connexin-43 expression as a the fetus such as fetal growth restriction and prerequisite for coordinated labor propaga- fetal (mis)programming with lifelong conse 

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quences, e.g. for later hypertension or stress regulation (Schleußner 2016a). The enzyme 11-beta-hydroxysteroid dehydrogenase (11ß-HSD), localized in the syncytiotrophoblast, catalyzes the reduction (11ß-HSD1) or oxidation (11ß-HSD2) of glucocorticoids. 11ß-HSD2 activity increases during pregnancy and converts active maternal cortisol to inactive cortisone, leaving the fetus unaffected by maternal cortisol levels. At term, the ratio of 11ß-HSD2:11ß-HSD1 activity increases, so that much more maternal cortisol enters the fetal circulation and contributes to the maturation of fetal adrenergic regulation (Wood and Keller-Wood 2016). In placental disorders with preeclampsia or fetal growth restriction, 11ß-HSD2 expression is reduced (Causevic and Mohaupt 2007). Synthetic glucocorticoids such as betaand dexamethasone or prednisolone are not or only slightly inactivated by 11ß-HSD, so that they have unrestricted effects on fetal stress regulation as well as fetal growth and maturation. This is used for the induction of fetal lung maturation with beta- or dexamethasone, but at the same time also leads to a significant placental and fetal growth restriction (Braun et al. 2015).

5.3

Peptide Hormones

5.3.1

Human Chorionic Gonadotropin (hCG)

hCG is the key essential embryonic hormone for the implantation of the embryo and the continuation of pregnancy. The embryo starts its hCG production already before implantation, so that hCG is detectable in the serum from day 8 post conception. During the first six weeks of gestation, embryonic hCG maintains the function of the maternal corpus luteum graviditatis and thus its progesterone and estrogen synthesis. Furthermore, it specifically induces decidual angiogenesis and

later myometrial tranquilization. Through the activation of dentritic cells, uNK and regulatory T cells and the inhibition of cytotoxic T cells and their cytokines, it contributes significantly to local immune modulation and immune tolerance at the fetomaternal interface (Nwabuobi et al. 2017). hCG is predominantly synthesized by the syncytiotrophoblast, but extravillous trophoblasts are also able to do so. The secretion, which mainly occurs into maternal blood, is pulsatile and doubles every 48 h until about week 10 of gestation, and then slowly decreases after this peak until the end of pregnancy (Costa 2016). hCG belongs to the family of glycoproteins with a similar structure as the pituitary hormones LH (luteinizing hormone), FSH (follicle-stimulating hormone) and TSH (thyroid-stimulating hormone). All these hormones consist of two subunits that are noncovalently linked—with identical α-subunit, the β-subunit differs specifically, which is why ßhCG is always measured, e.g. as a pregnancy test. However, ßhCG is also 80–85% homologous to the LH ß-subunit. The ßhCG gene cluster is located on chromosome 19. hCG activates adenylate cyclase, ­phospholipase C and ion channels, which in turn control intracellular cAMP, inositol phosphate and Ca2+ and regulate a variety of intracellular signaling cascades, such as e.g. protein kinase A and C, ERK1/2MAPK or Smad2 through binding to the G-protein coupled LH/hCG gonadotropin receptor or through direct and indirect interaction with the TGFß receptor (Nwabuobi et  al. 2017). cAMP, via protein kinase A, causes cytotrophoblast cell fusion and microvillous formation, which is essential for syncytiotrophoblast development. At the same time, hCG/LH receptors are expressed in the cytotrophoblast, syncytiotrophoblast, and also the extravillous trophoblast, so that hCG also regulates trophoblast invasion and exerts local autocrine and paracrine effects (Handschuh et al. 2007).

99 Endocrinology of the Placenta

hCG systemically alters maternal endocrine homeostasis. The pituitary-ovarian axis is suppressed by negative feedback regulation of hCG at the LH receptor of the adenohypophysis, resulting in ovarian dormancy during pregnancy (Schleußner 2017). hCG also binds to the TSH receptors and thus stimulates thyroid function, whereas pituitary TSH secretion is slightly reduced in the first trimester of pregnancy. Excessively high hCG levels, e.g. in the case of a placental tumor (hydatidiform mole), can therefore trigger a full-blown thyrotoxic crisis. In the second half of pregnancy, TSH levels rise to maintain the physiological euthyroidism of pregnancy with intact pituitary-­ thyroid regulation. The essential mechanism for this is the doubling of estrogen-­ induced hepatic synthesis of thyroxine-­ binding globulin (TBG). Thus, with free thyroxine (T4) and triiodothyronine (T3) levels remaining constant, total T3 and T4 concentrations increase in the first half of pregnancy to remain stable after 20 weeks of gestation (Ross 2017). 5.3.2

Leptin

Leptin plays a central role in the regulation of energy homeostasis, appetite and body weight, but also influences angiogenesis, immune regulation and reproduction (Park and Ahima 2015). Mainly produced in adipocytes, the placenta synthesizes large amounts of leptin during pregnancy, which is secreted into the maternal and fetal circulation as well as into the amniotic fluid. During early pregnancy, leptin synthesis is highest in the syncytiotrophoblast. Maternal leptin levels increase throughout pregnancy until the beginning of the 3rd trimester and then remain stably high until birth (Costa 2016). With permanently elevated leptin levels, the mother is in a physiological leptin

5

resistance, so that despite high leptin concentration, its central satiety effect is absent, resulting in hyperphagia and increased weight gain as well as hyperinsulinemia. This condition is accompanied by increased lipolysis, so that the mother can primarily use the lipids, which are only available to a limited extent in the placenta, as an energy source, while the glucose reserves are available for fetal supply. This can also be interpreted as a physiological adaptation to the increasing supply requirements of the growing fetus and as a mobilization of the maternal energy reserves (Ladyman et al. 2010). At the same time, leptin plays an essential role in fetal growth control, angiogenesis and hematopoiesis. Within the placenta it has paracrine functions and has an immunomodulatory effect (. Fig. 5.2) (Ashworth et al. 2000). Leptin binds to the membranous leptin receptor (LepR), which is present ubiquitously in the human organism in six different splice variants and can activate JAK-STAT, ERK1/2 and PI3K signaling cascades. Leptin’s paracrine effect is mediated by LepR in the syncytiotrophoblast, where it also influences the syncytial hormone synthesis, for example. It increases hCG production while inhibiting estradiol and progesterone synthesis (Costa 2016). In the extravillous trophoblast, leptin has a proliferative effect and is involved in the regulation of invasion via expression of metalloproteinases. Leptin plays a key role in local immunomodulation by interfering with the regulation of prostaglandin and inflammatory cytokines, modulating uterine NK cell activity and stimulating trophoblast HLA-G expression (Pérez-Pérez et al. 2015). At the same time, it plays an important local angiogenic role in decidual neovascularization and uterine vascular remodeling by enhancing vascular endothelial growth factor (VEGF) expression.  

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Maternal Placental Fetal

5

Plasma leptin concentration

Adipose tissue Delivery

ObRb ObRa ObRe

Gestational age

Fetal leptin (adipose tissue and other tissues)

Leptin-binding protein Possible functions of leptin Maternal body: Mobilisation of energy reserves

Placenta: Growth, Angiogenesis, Immunomodulation

Fetus: Growth, Aangiogenesis, Hematopoiesis, Brain development, Immunity

..      Fig. 5.2  Leptin as a placental regulator of maternal and fetal supply. (From Schleußner 2017, modified from Ashworth et al. 2000). ObR Obesity Receptor

5.3.3

Corticotrophin Releasing Hormone (CRH)

CRH is a 162 amino acid peptide hormone synthesized in the hypothalamus during pregnancy, but predominantly in the syncytiotrophoblast of the placenta and secreted into both the maternal and fetal circulation. Placental CRH plays a central role in physiological labor induction by coordinating fetal and maternal endocrine signals and activating the molecular mechanisms of labor induction (. Fig.  5.3) (Schleußner 2016b). CRH levels increase exponentially during pregnancy, and a correlation of concentrations already measured in the 1st trimester with the time of birth (preterm or term delivery, postterm pregnancy) was found. In preterm labor, significantly increased plasma levels are found compared to stable  

pregnancies of the same gestational age, (Petraglia et al. 2010). Placental, as well as hypothalamic CRH synthesis, is modulated by neurotransmitters and neuropeptides such as vasopressin. In contrast to the regulation of the adrenergic hypothalamic–pituitary axis, placental CRH synthesis is subject to positive feedback from cortisol, which originates either from the fetal adrenal cortex or from the mother. This may also explain the labor-­ inducing effects of acute maternal or fetal stress. The administration of glucocorticoids for lung maturation also causes a transient increase in CRH, which can also lead to a temporary increase in uterine contractions (Schleussner et al. 2000). Progesterone, on the other hand, inhibits trophoblast CRH synthesis. The antagonism of the two steroid hormones is of central importance in CRH regulation. While the

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101 Endocrinology of the Placenta

Fetal membranes and placenta

Fetus

Possible negative feedback

Mother

Cortisol

Cortisol

Hypothalamus

Hypothalamus Cholesterol

CRH

∆5 -pregnenolone

Anterior Pituitary

Cortisone Progesterone

Posterior Pituitary

Dehydroepiandrosterone

Corticotropin

Oxytocin

Estriol

+

Adrenal gland

+

+

Oxytocin from the placenta

+

+

Prostaglandin E2 Prostaglandin F2α

+ +

+

CRH from the placenta

+

DHEAS Cortisol Preparation of the fetal organ systems for delivery

Positive feedback

Prostaglandin receptors Oxytocin receptors Gap junctions Uterus

Rupture of fetal membranes

Labour

..      Fig. 5.3  Endocrine regulation of labor. (From Schleußner 2010, modified after Challis 2001). CRH Corticotrophin Releasing Hormone; DHEAS Dehydroepiandrosterone

inhibitory effect of progesterone predominates during pregnancy, increasing cortisol production by the fetal adrenal cortex towards the end of pregnancy leads to increased placental CRH secretion (Smith 2007). The progressive CRH increase in turn leads to activation of the fetal adrenergic axis and thus also stimulates the secretion of fetal DHEAS, which is aromatized to estrogen in the placenta. As described in 7 Sect. 5.2.2, this local estrogen dominance increases oxytocin receptor expression, gap junction formation and prostaglandin synthesis in the decidua, chorion and amnion. Prostaglandin synthesis enzymes (PGHS-2) are activated by increasing CRH levels, while at the same time  

the degradation of prostaglandin by prostaglandin dehydrogenases is inhibited. Thus, there is a direct effect of cortisol on both prostaglandin synthesis capacity and inhibition of prostaglandin metabolism (Challis 2001). Independently, direct effects of placental CRH on uterine contractility via myometrial CRH receptors are also found. In addition to CRH, urocortins are also produced in the placenta, amnion and decidua, which belong to the CRH family and show up to 45% structural homology with CRH.  Like CRH, urocortins act in stress regulation and birth induction, but also control endothelial vascular tone in uteroplacental perfusion (Petraglia et al. 2010).

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5.3.4

 lacental Lactogen (hPL) P and Placental Growth Hormone (hPGH)

Human placental lactogen (hPL), as well as human placental growth hormone (hPGH) are peptide hormones, which show a large structural homology and are encoded in the same gene cluster on chromosome 17, whereby hPL functionally acts more like prolactin (Handwerger and Freemark 2000). Both hormones are synthesized mainly in the syncytiotrophoblast, but also in the extravillous trophoblast. hPL is detectable as early as two weeks post conception, hPGH from the 3rd week onwards. The levels of both hormones rise sharply over the course of pregnancy (hPL more than hPGH), while hypothalamic– pituitary GHRH and GH secretion are suppressed. Thus, placental lactogens and growth hormones take over control of both the maternal and fetal GH-IGF axes (. Fig. 5.4) (Newbern and Freemark 2011). Maternal hPL and hPGH blood levels correlate with placental size and are regulated by blood glucose concentration. Starvation and hypoglycemia stimulate  

hPGH

hPGH secretion, while glucose administration inhibits hPGH secretion in  vitro and in  vivo. Unlike pituitary GH, hPGH is tonically secreted and not subject to control by GHRH, but is enhanced by PPAR-γ and cAMP and inhibited by leptin, insulin and cortisol. hPL, on the other hand, is stimulated by GHRH, insulin, apolipoproteins, PPAR-γ and cAMP (Costa 2016). Both hormones bind to the ubiquitous membrane-bound GH receptor, although hPL has a low binding affinity, while it can also bind to the prolactin receptor. hPL enhances maternal lipolysis so that the resulting free fatty acids are available as an energy source. hPGH acts as an insulin antagonist and influences maternal glucose metabolism to ensure that maternal energy stores can be used for a constant supply to the fetus. It induces maternal IGF-1 synthesis, which contributes to genital organ growth. It also enhances uteroplacental perfusion and, in turn, fetoplacental supply. Thus, in a coordinated interplay of the placental hormones leptin, hPL and hPGh, maternal metabolism is tuned to the needs

Maternal IGF-1 Estrogen

Maternal insulin resistance

Uterine growth Placental perfusion

Fetal growth

GDM risk Glucocorticoids Inflammatory cytokines Progesterone

Beta cell expansion

Insulin

PRL, hPL ..      Fig. 5.4  Effects of placental lactogen (hPL) and placental growth hormone (hPGH) on the regulation of maternal metabolism and fetal growth and on the

risk of gestational diabetes. (Modified from Newbern and Freemark 2011). GDM Gestational diabetes, IGF-1 Insulin-like growth factor 1, PRL Prolactin

103 Endocrinology of the Placenta

of fetal development (. Fig. 5.4) (Newbern and Freemark 2011). hPL is secreted into both fetal and maternal circulations, but hPGH is secreted only into maternal blood, so that its effects are directed exclusively to the maternal metabolism. hPL, on the other hand, directly affects fetal growth by enhancing IGF-1 synthesis in the fetal liver and pancreas. Paracrine local effects of these hormones are still poorly understood, but since growth hormone receptors are also present in the placenta, these hormones have been demonstrated for placental growth and also trophoblast invasion. A number of other growth factors are also secreted in the placenta, such as somatostatin, ghrelin, adiponectin, IGF-1 and IGF-2, which both regulate the GH-IGF axis and thus are the effectors of this axis (Petraglia et al. 1996).  

5.3.5

Summary

The placenta is the largest endocrine organ during pregnancy dominating the entire hormonal system. Placental peptide and steroid hormones influence all maternal endocrine functional circuits and regulatory axes. Progesterone, as the central pregnancy-­ maintaining hormone, relaxes the myometrium and has anti-inflammatory and immunosuppressive functions that allow the formation of the necessary immune tolerance at the fetomaternal interface. Estrogens regulate maternal adaptation to pregnancy. Leptin, placental lactogen, and placental growth hormone alter maternal metabolism to use its resources to provide continuous fetal nutrition. Placental corticotrophin releasing hormone (CRH) is central to the initiation of labor as the placental clock.

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References Ashworth C, Hoggard N, Thomas L, Mercer JG, Wallace JM, Lea RG (2000) Placental leptin. Rev Reprod 5:18–24 Bjornstrom L, Sjoberg M (2005) Mechanisms of estrogen receptor signaling: convergence of genomic and nongenomic actions on target genes. Mol Endocrinol 19:833–842 Braun T, Sloboda DM, Tutschek B, Harder T, Challis JR, Dudenhausen JW, Plagemann A, Henrich W (2015) Fetal and neonatal outcomes after term and preterm delivery following betamethasone administration. Int J Gynaecol Obstet 130(1): 64–69 Bukovsky A, Cekanova M, Caudle MR, Wimalasena J, Foster JS, Henley DC, Elder RF (2003) Expression and localization of estrogen receptoralpha protein in normal and abnormal term placentae and stimulation of trophoblast differentiation by estradiol. Reprod Biol Endocrinol 1:13 Causevic M, Mohaupt M (2007) 11beta-­ Hydroxysteroid dehydrogenase type 2  in pregnancy and preeclampsia. Mol Aspects Med 28(2):220–226 Challis JR (2001) Understanding pre-term birth. Clin Invest Med 24:60–67 Chang K, Zhang L (2008) Steroid hormones and uterine vascular adaptation to pregnancy. Reprod Sci 15(4):336–348 Corcoran JJ, Nicholson C, Sweeney M, Charnock JC, Robson SC, Westwood M, Taggart MJ (2014) Human uterine and placental arteries exhibit tissue-­ specific acute responses to 17β-estradiol and estrogen-receptor-specific agonists. Mol Hum Reprod 20(5):433–441 Costa MA (2016) The endocrine function of human placenta: an overview. Reprod Biomed Online 32(1):14–43 Csapo AI, Knobil E, van der Molen HJ, Wiest WG (1971) Peripheral plasma progesterone levels during human pregnancy and labor. Am J Obstet Gynecol 110:630–632 Diczfalusy E (1984) The early history of estriol. J Steroid Biochem 4:945–953 Di WL, Lachelin GC, McGarrigle HH, Thomas NS, Becker DL (2001) Oestriol and oestradiol increase cell to cell communication and connexin43 protein expression in human myometrium. Mol Hum Reprod 7:671–679

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Goldman S, Shalev E (2007) Progesterone receptor profile in the decidua and fetal membrane. Front Biosci 12:634–648 Groothuis PG, Dassen HH, Romano A, Punyadeera C (2007) Estrogen and the endometrium: lessons learned from gene expression profiling in rodents and human. Hum Reprod Update 13:405–417 Halasz M, Szekeres-Bartho J (2013) The role of progesterone in implantation and trophoblast invasion. J Reprod Immunol 97:43–50 Handwerger S, Freemark M (2000) The roles of placental growth hormone and placental lactogen in the regulation of human fetal growth and development. J Pediatr Endocrinol Metab 13:343–356 Handschuh K, Guibourdenche J, Tsatsaris V, Guesnon M, Laurendeau I, Evain-Brion D, Fournier T (2007) Human chorionic gonadotropin expression in human trophoblasts from early placenta: comparative study between villous and extra villous trophoblastic cells. Placenta 28:175–184 Kuon RJ, Abele H, Berger R, Garnier Y, Maul H, Schleußner E, Rath W (2015) Progesterone for Prevention of Preterm Birth—Evidence-based Indications. Z Geburtshilfe Neonatol 219(3): 125–135 Ladyman SR, Augustine RA, Grattan DR (2010) Hormone interactions regulating energy balance during pregnancy. J Neuroendocrinol 22:805–817 Newbern D, Freemark M (2011) Placental hormones and the control of maternal metabolism and fetal growth. Curr Opin Endocrinol Diabetes Obes 18:409–416 Nwabuobi C, Arlier S, Schatz F, Guzeloglu-Kayisli O, Lockwood CJ, Kayisli UA (2017) hCG: biological functions and clinical applications. Int J Mol Sci 18(10):E2037. https://doi.org/10.3390/ijms18102037 Park HK, Ahima RS (2015) Physiology of leptin: energy homeostasis, neuroendocrine function and metabolism. Metabolism 64:24–34 Pérez-Pérez A, Sánchez-Jiménez F, Maymó J, Dueñas JL, Varone C, Sánchez-Margalet V (2015) Role of leptin in female reproduction. Clin Chem Lab Med 53(1):15–28 Petraglia F, Florio P, Nappi C, Genazzani AR (1996) Peptide signaling in human placenta and ­membranes: autocrine, paracrine, and endocrine mechanisms. Endocr Rev 17(2):156–186

Petraglia F, Imperatore A, Challis JR (2010) Neuroendocrine mechanisms in pregnancy and parturition. Endocr Rev 31(6):783–816 Renthal N, Koriand’r CW, Montalbano AP, Chien-­ Cheng C, Lu G, Mendelson C (2015) Molecular regulation of parturition: a myometrial perspective. Cold Spring Harb Perspect Med 5:a023069 Ross DS (2017) Overview of thyroid disease in pregnancy. In Cooper DS, Lockwood CJ, editors. UpToDate, http://www.­uptodate.­com/contents/ overview-­o f-­t hyroid-­d isease-­i n-­p regnancy. Accessed on 30 Nov 2017 Schleußner E (2010) Normale Geburt. In: Kiechle M (Hrsg) Gynäkologie und Geburtshilfe, 2. Aufl. Elsevier, München, S 379–388 Schleußner E (2016a) Fetale programmierung. In: Schneider H, Husslein P, Schneider KTM (Hrsg) Die Geburtshilfe, 5. Aufl. Springer, Heidelberg, S 367–378 Schleußner E (2016b) Molekulare Grundlagen von Wehentätigkeit und Geburt. In: Liedtke C, Rody A (Hrsg) Molekulare Gynäkologie und Geburtshilfe für die Praxis, 5. Aufl. Thieme, Stuttgart, S 42–46 Schleußner E (2017) Physiologische Veränderungen in der Schwangerschaft. In: Fischer-Betz R, Østensen M (Hrsg) Rheumatische Erkrankungen in der Schwangerschaft. De Gruyter Berlin, Boston S, S 11–26 Schleussner E, Weitschat T, Hüller M, Albrecht S, Möller U, Seewald H-J (2000) Effects of glucocorticoid administration on placental CRH secretion and fetal and maternal adrenal function in  vivo. Placenta 21:42–43 Smith R (2007) Parturition. N Engl J Med 356: 271–283 Toriola AT, Vääräsmäki M, Lehtinen M et  al (2011) Determinants of maternal sex steroids during the first half of pregnancy. Obstet Gynecol 118: 1029–1036 Tuckey RC (2005) Progesterone synthesis by the human placenta. Placenta 26:273–2281 Wood CE, Keller-Wood M (2016) The critical importance of the fetal hypothalamus-pituitary-adrenal axis. F1000Research 5(F1000 Faculty Rev):115. https://doi.org/10.12688/f1000research.7224.1

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Teratology Herbert Juch Contents 6.1

Introduction – 106

6.2

Congenital Anomalies Historically – 106

6.3

Congenital Anomalies Today – 108

6.4

Basic Risk – 109

6.5

Medication and Pregnancy – 110

6.6

Placenta and Teratology – 112

6.6.1

T he Sensitivity of the Embryo to Toxic Influences Depends on the Genotype – 112 The Sensitivity of the Embryo to Toxic Influences Depends on Its Stage of Development – 114 Different Embryotoxic Influences Affect (Embryonic) Development via Relatively Few Specific Mechanisms – 116 After Exposure to Teratogens, Different Developmental Courses Are Possible in Principle – 116 The Way in Which Toxic Influences Reach the Embryo/Fetus Depends on Their Physical and Chemical Properties – 117 Dose-response Relationships Apply in Teratology as Elsewhere in Pharmacology and Toxicology – 118

6.6.2 6.6.3 6.6.4 6.6.5 6.6.6

6.7

Conclusion – 120 References – 120

© Springer-Verlag GmbH Germany, part of Springer Nature 2023 B. Huppertz, E. Schleußner (eds.), The Placenta, https://doi.org/10.1007/978-3-662-66256-4_6

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6.1

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Introduction

Teratology is considered to be the science of “congenital anomalies”, or congenital “malformations” or “birth defects”. Originally, teratology was primarily morphologically oriented and a distinction was made in German between malformations and disruptions. In the case of malformations, an endogenous (genetic) cause was assumed for the congenital morphological peculiarities; in the case of disruptions, an exogenous cause was assumed as a disturbance of the development, which would otherwise have proceeded normally. In the meantime, the term “disruption” is no longer particularly common, and “malformation” is also frequently used for exogenously caused shape abnormalities. In the teratological context, it makes sense to use the term “congenital anomaly” in order not to be limited to congenital abnormalities in the shape of the child, but also to include functional abnormalities. “Anomaly” is to be understood as a deviation from the norm, which has disease value, thus requiring therapeutic intervention. In Anglo-American usage, a distinction is often made between “major congenital anomalies” and “minor congenital anomalies”, the latter being characterized by the fact that they do not represent functional or cosmetic impairments requiring medical treatment. In German, the term “große Fehlbildung” (major malformation) is occasionally used in reference to this. It should be noted that there are regular discussions in specialist circles about the classification of anomalies as “major” or “minor”, as this classification can influence the assessment of teratological risks in the scientific literature. >>Currently, the focus of teratological interest is not only on infantile organ or skeletal anomalies, but also on miscarriage and prematurity, placental dysfunction, intrauterine malnutrition and,

in addition, prenatal influences and their possible long-term consequences for social and intelligence development or for disease dispositions.

Thus, a very broad field of activity has arisen for teratology in the meantime, motivated by the clear teratological mandate to recognize various exogenous negative influences on human prenatal development as such and to protect pregnant women from them as far as possible. In view of the wide range of tasks, it is hardly surprising that teratology today is more than ever an interdisciplinary science, with human genetics, gynecology and obstetrics, pediatrics, pharmacology and toxicology, epidemiology, hygiene and infectiology, occupational medicine and social medicine being particularly involved, in addition to embryology including placenta research, and each discipline has naturally developed its own approaches to the subject and its own focal points.

6.2

Congenital Anomalies Historically

Ever since there have been medical records, great attention has been paid to the morphological phenomenon of “miracle formations”, from whose Greek term τέρας téras (miracle thing, omen, also monster) the term teratology is ultimately derived. The birth of a child with sometimes frightening, scary, mystically interpreted as “signs”, was given great importance not only by the parents concerned, but also by society. There has always been a need to find the causes of these problems, which are experienced as fateful. In what we now consider “pre-­scientific” times, concepts such as sin and guilt, as well as religious worldviews, played a central role in finding and combating the causes of congenital anomalies (cf. Schumacher et al. 1992). In part, however, quite useful explanations for observed health problems were already pos-

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107 Teratology

tulated in antiquity, such as the trauma theory, the spatial theory, or the disease theory (see the following overview). Nevertheless, not least Christian religious interpretations of the subject sometimes had life-threatening consequences for affected children, their parents, but also, objectively speaking, completely uninvolved third parties (“witches”, “sorcerers”, possibly “evil beggars”). Attempts to Explain Anomalies, Historically

Congenital

55 Trauma theory: e.g. blow to the abdomen of a pregnant woman 55 Space theory: a too narrow uterus/too much semen 55 Theory of disease, heredity: diseases of the fetus, familial clustering 55 Lack of semen or “maternal matter” 55 Rotten semen 55 Woman’s “Versehen”: mother’s imagination triggers malformations

55 Poor posture of the pregnant woman 55 Sins committed: e.g. “illegitimate” sexual intercourse 55 Human-animal intercourse 55 Devils, trolls, evil spirits 55 Trickery of evil beggars (often people with congenital anomalies) 55 God wants to show us his power According to Schumacher et  al. (1992) and Palister and Paré (1982).

>>The search for the causes of congenital anomalies has lost none of its explosive power to this day. The prenatal and postnatal medical-diagnostic effort expended for this purpose is now considerable. Nevertheless, in about two out of three affected children, no clear cause for the congenital health problems can be identified (. Table 6.1).  

.       Table 6.1  Causes of congenital anomalies, natural scientific Causes

Examples

Share (%)

Genetic

Monogenic diseases, chromosomal anomalies

~11–30

Chemical/physical

Drugs, medication, pollutants, hyperthermia, hypoxia, ionizing radiation

~2–4

Maternal disease

Hypothyroidism, diabetes mellitus, gestoses, phenylketonuria, severe obesity, systemic lupus erythematosus (SLE), myasthenia, etc. Infections: Cytomegalovirus, rubella, Erythema infectiosum (Fifth disease), varicella, listeriosis, syphilis, toxoplasmosis, zika virus

~3

Anatomical

Twins, oligohydramnios, uterine anomalies

~3

Unclear

Polygenic or multifactorial anomalies, different combinations of endogenous and exogenous causes

~65–80

Adapted from Schäfer et al. (2012), Rösch and Steinbicker (2003), Schardein (2000), Enders (1991), and Wilson (1977)

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6.3

H. Juch

Congenital Anomalies Today

rising insurance rates for the corresponding professional liability insurance policies).

Of course, the perception of congenital anomalies has significantly changed and objectified by the science-based, biopsychosocial approach. Today, congenital anomalies appear to us as a multi-layered problem, whereby various causally relevant levels hold important starting points both diagnostically and therapeutically (. Fig.  6.1). The old question of guilt has, however, been preserved—albeit sometimes with a focus on the subconsciousness of those affected. It finds its contemporary expression, on the one hand, as a feeling of guilt and self-­ incrimination on the part of the parents concerned and, on the other hand, as an intensive legal search for guilt, e.g. on the part of medical doctors and midwives who accompany pregnancy and childbirth (which can also be clearly seen in their constantly

The lack of a clear cause for the congenital anomalies of a child in about two out of three cases is perceived by most of the affected parents as particularly stressful and offers especially today, in the “post-factual” internet age, a lot of (virtual) space for various irrational to abstruse attempts at explanation. Therefore, a high degree of sensitivity, but also of clarity, is necessary from the professional side in the care of children with congenital anomalies and their parents, in order to avoid at least the errors and fallacies that are nowadays clearly identified as such.

of the preg uation nan l sit a tw i c om so o an h e h p t regna c f o y y s nt log o w i om ys ph Placenta an

Bi o

6

P

 

Tip

genetics Epi Paternal Maternal

Genome

..      Fig. 6.1  Diagnostically and therapeutically relevant levels in connection with congenital anomalies. (Modified after Schumacher et al. 1992)

109 Teratology

6.4

Basic Risk

Important for the understanding of congenital anomalies and their potential avoidability is the knowledge of a fundamental imperfection of all natural beings and developments—including the prenatal development of humans as embryos or fetuses. This finds expression in the so-­called basic risk of congenital anomalies, i.e. a natural error rate or a lack of precision in our prenatal development. Even if, by all accounts, an ideal “healthy” pregnant woman (according to the WHO definition: “Health is a state of complete physical, mental and social well-being …”) can be assumed to have an optimal course of pregnancy, without any family history of genetic diseases and without any recognizable external negative influences, congenital anomalies occasionally occur. >>This baseline risk of congenital (i.e., not postnatally induced) health problems in humans in the 1st year of life is estimated to be ~3% for “normal” average healthy pregnant women, with normal average pregnancy outcomes. In addition, a ~15% “natural” risk of miscarriage is observed in the 1st trimester.

However, this risk of miscarriage increases markedly with increasing maternal age (and to a lesser extent with increasing paternal age) and thus indicates, among other things, a connection with the statistically more frequent age-related stochastic chromosomal abnormalities in oocytes (as well as in sperm cells). In addition, a certain proportion of these abortions can be attributed to pathological placentation (occasionally also immunologically caused), a direct indication of the placental influence on the basic reproductive risks. These natural, undesirable events can sometimes be explained by random newly occurring dominant or familial recessive mutations in the germ cell genome. These

6

events are also ultimately understandable in terms of systems theory and are due to the complexity of the system “prenatal human development” with its highly interconnected interactions of all genetic, epigenetic and exogenous factors. Similar to other highly complex systems, even with theoretical knowledge of all relevant measurands at the beginning of human development, the outcome of development could not be exactly  predicted. Therefore, for embryonic and fetal development, sensitivity to developmental perturbations must be postulated through “weak causalities.” Weak causality means, tiny changes in certain initial conditions, minimal fluctuations in physiologic parameters, which can lead to recognizable developmental differences, up to and including congenital anomalies, in the further course of development. In weather forecasting, this phenomenon is known as the “butterfly effect” (Lorenz 1972), where subtle air movements, as caused by butterflies, may be responsible for the development of a subsequent storm in a distant area. Different organ systems may well have different sensitivities in this respect, which would explain why the baseline risk for certain anomalies (e.g. heart malformations) is much higher than for others. Various known risk factors increase these baseline reproductive risks to varying degrees, although usually no factor causes a 100% risk of abnormality. >>From the concept of “baseline risk” it can be deduced that no pregnant woman has a guarantee of a healthy baby, at the same time, generally harmful influences will not  lead to health problems in the baby with absolute certainty.

An important disadvantage of this statistically based approach, which reflects reality very well, is the impossibility of accurately predicting any congenital problems in individual cases. Prospectively, only probability statements can be made. In addition, retro-

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spectively, if congenital anomalies are present in the child, the causality cannot be clarified exactly in many cases. For example, after a pregnant woman’s one-time excessive alcohol consumption (“binge drinking”) between the 4th and 8th week of embryonic development, an increased risk of congenital anomalies in the child is to be expected. The question of the concerned woman, whether the child will be damaged after this unintentional exposure to alcohol in early pregnancy or not, cannot be answered clearly with “yes” or “no”. It could be stated that the chances of having a  normal  baby would probably be well over 90%. However, the chances would be statistically somewhat lower than they would have been without this incident. If, for example, a heart defect were to be diagnosed in the child after such an exposure, i.e. an anomaly whose basic risk is just under 1% and which we know is caused “multifactorially” (i.e. by various genetic factors in combination with various environmental factors), no clear causality of alcohol consumption could be established. However, a possible contribution of the undisputed teratogenic noxious agent alcohol could not be clearly excluded either.

6.5

Medication and Pregnancy

One aspect of teratology regularly receives special attention in the clinical field: the use of medication during pregnancy. This topic was strongly influenced by the thalidomide disaster at the end of the 1950s and beginning of the 1960s. Thalidomide, which was initially considered harmless and relatively non-toxic, turned out to be one of the most potent teratogenic substances in humans. Only after thousands of children had been born with sometimes severe limb malformations was it possible to identify thalidomide

ingestion during pregnancy as the cause. As a consequence of this teratological worst case scenario, efforts in the field of reproductive toxicology were significantly intensified. For example, the insufficiency of reproductive toxicology testing of medications prior to marketing approval in only one animal species was recognized and mandatory teratological testing of agents in at least a second species was introduced. Teratological research on negative exogenous influences on prenatal development has generally received a strong boost in the wake of this medical scandal. >>As a negative after-effect of the thalidomide catastrophe, a partly undifferentiated and often exaggerated, frequently also completely unjustified concern about the use of medications during pregnancy has remained to this day.

This concern affects not only the pregnant women themselves, but not infrequently also their attending physicians. In line with the spirit of the times of an increasingly anxious society that is striving more and more for security, attempts are made in this area to “protect” themselves legally against possible lawsuits based on congenital anomalies in children by means of sophisticated non-binding formulations, and a primarily legal ethic is imposed on the medical doctor-patient relationship. The pregnant woman then easily gets the impression that she is not regarded as a responsible partner in the attempt to resolve a health problem, but as a client and potential opponent in court. This results in a loss of trust in the patient, and instead of facilitating a discerning risk-benefit assessment of a medication therapy during pregnancy, it occasionally leads to wrong decisions  with negative long-term consequences.

111 Teratology

Tip

To avoid such undeniably disadvantageous developments for both medical doctors and patients in the question of taking medication during pregnancy, the principle of “shared decision  making” would be appropriate (Elwyn et  al. 2012). This involves the patient to the maximum extent possible in the complex decision-making process.

Through optimal information about the opportunities and risks of the available therapeutic alternatives, the pregnant woman should be put in a position to decide on one of the possible therapeutic options (including “non-treatment”) according to her individual needs and preferences, together with the attending physician. The prerequisite for such “informed decision-making” on the use of medication during pregnancy is, in addition to a realistic assessment of the performance of the therapeutic options, a realistic assessment of any teratological risks.

the past decades, which must of course be continuously expanded to enable a wellfounded risk assessment in the future even for active substances that are currently little studied. For individual teratological risk assessment, in addition to knowledge of the currently available data from observational studies on the use of specific medication in humans and of the available animal repro-­ tox data, knowledge of the “basic rules” or “principles” of teratology established as early as the 1970s by James Wilson (Friedman 2010) has proved helpful (see following overview), which have only been slightly modified in the meantime. Based on fundamental findings on embryonic and fetal development from animal experiments and observations on human embryology, they facilitate both the individual optimization of therapy in the ideal case of therapy planning before the onset of pregnancy and the realistic risk assessment of accidental substance exposure in the case of unplanned pregnancy. Wilsonʼs Rules (Modified After Wilson 1977)

Tip Support for individual risk assessment is provided by teratology information services (TISes, 7 https://www.­entis-­org.­eu; https://mothertobaby.org) set up worldwide.  

These free-of-charge counselling facilities are also endeavoring to expand knowledge of teratogenic medication effects in humans by means of follow-up studies on a continuous and internationally networked basis. Through these activities, but also through various other teratological and pharmacoepidemiological studies, e.g. based on malformation and drug intake registries, a very comprehensive knowledge of the safety and risks of individual active substances in human pregnancy has been acquired over

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1. The sensitivity of the embryo to toxic influences depends on its genotype 55 Different medication effects in humans and experimental animals (species specificity of teratogenic effects) 55 Sensitivity to teratogens can vary from person to person (enzyme polymorphisms) 2. The sensitivity of the embryo to toxic influences depends on its developmental stage 55 Abortion or regeneration (“all or none” phenomenon) in the 1st and 2nd week of development 55 In the first trimester mostly severe morphological and functional anomalies (embryopathies)

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55 In the second and third trimesters, mostly mild, predominantly functional anomalies (fetopathies) 55 Specific teratogenic noxious agents often only have an effect at certain times in development 3. Different embryotoxic influences affect (embryonic) development via relatively few specific mechanisms 55 Spectrum of possible congenital anomalies is limited 55 Various harmful influences or genetic causes cause similar abnormalities 55 A teratogenic effect does not increase the risk of all possible congenital anomalies 4. After exposure to teratogens, the following developmental courses are possible in principle 55 Normal development, defects are completely repaired (especially 1st and 2nd week of development) 55 Intrauterine fetal death, miscarriage, premature birth 55 Morphological organ anomalies, functional limitations, growth restriction 55 Tumor development 5. The way toxic influences reach the embryo/fetus depends on their physical and chemical properties 55 Pharmacokinetics in pregnancy: –– Medication kinetics in the mother –– Medication kinetics in placenta/embryo/fetus 6. In teratology, dose-response relationships apply as elsewhere in pharmacology and toxicology 55 Minimal teratogenic dose thresholds for teratogenic effects (NOAEL, “no observed adverse effect level”) 55 Lower dose, lower risk of damage with exposure above threshold

6.6

Placenta and Teratology

>> The placenta plays a central role, especially in the context of exogenously influenced and co-induced congenital anomalies, on the one hand due to its complex barrier function for diverse exogenous influences during prenatal development and on the other hand as a direct target organ for teratogenic damage with consecutive placental pathologies (7 Chaps. 7, 8, 10, 11) and secondary pregnancy complications as well as health problems in the child.  

To shed light on the role of the placenta from the point of view of teratology, an attempt will now be made to illustrate its significance on the basis of the basic teratological rules. 6.6.1

The Sensitivity of the Embryo to Toxic Influences Depends on the Genotype

A classical derivation from this basic rule refers to the limited applicability of reproductive toxicological data obtained in animal experiments to the human situation. >>The fundamental species differences in the genetic blueprint of mammals are particularly evident phenotypically in the very different strategies of placentation.

Macroscopically and tissue architecturally, interesting variations can be observed in this regard. From a functional point of view, both with regard to the organization of the perfusion of the organ and with regard to the invasive behavior of the placenta and the associated different challenges for the maternal immune system (tolerance, invasion control), there are major differences between

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species. In terms of embryonic toxicology, these genetically determined differences are of particular importance when attempting to apply the correlations between blood levels of potentially harmful substances and their concentration in the embryonic and fetal compartments, which have been established in common animal models (mouse, rat, rabbit), to the situation in humans. In the case of readily absorbable lipophilic small molecules, which also efficiently cross other tissue barriers such as the blood-brain barrier and are distributed more or less over all body compartments anyway, smaller species differences in embryonic or fetal exposure are more likely to be expected. In contrast, this may be much more differentiated in the case of larger molecules, such as the biologicals  that are already being used more and more frequently (see below), or nanoparticles that may be used in the future for diagnostic or therapeutic purposes. In any case, genetically determined species-­specific differences in the metabolism of the tissues involved in the placental barrier, in particular the trophoblast cell layers, but also the endothelial cells of the fetal vessels, must be taken into account for lipophilic small molecules. Depending on their transporter and enzyme equipment or activity, the exposure of the embryo or fetus may vary considerably at a given maternal blood level (see 7 Sect. 6.6.5 “The way in which toxic influences reach the embryo/fetus depends on their physical and chemical properties”). However, this first basic rule also points to possible differences in sensitivity to teratogenic noxae within a species, which can be observed even with absolutely identical exposure due to identical maternal blood levels. This was impressively demonstrated in a dizygotic and heteropaternal pregnancy (two oocytes, two different fathers) of an epilepsy patient who had been treated with phenytoin (Phelan et al. 1982). One twin was

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diagnosed with a “fetal hydantoin syndrome” with multiple congenital anomalies, while the second child was completely normal. Genetically different epoxide hydroxylase (EPHX1) enzyme activity was identified as the presumptive cause. The affected child showed low activity of this enzyme responsible for the degradation of toxic phenytoin metabolites, while the unaffected child showed high activity. EPHX1 appears to be expressed to a greater or lesser extent by almost all cells in the body however, there is currently no knowledge of the extent of expression in the early placenta, so that a possible placental contribution to the described discordance of the twin children cannot be estimated. Nevertheless, initial work on a defined placenta-specific pharmacogenetic contribution to fetal exposure is already available, e.g. in the context of known genetic variants of transport proteins from the group of multi-drug resistance proteins such as ABCB1 (Bliek et  al. 2009). The plausible background for these considerations is that certain alleles of such transporters are associated with a reduced function of these “detoxification molecules” expressed in the placenta. Expectant mothers (and fathers) who can pass on such genetic variants with reduced activity to their offspring would have a higher risk of congenital anomalies in the child in the event of foreign substance (e.g. medication) exposure during pregnancy. However, it would be important to know the actual genotype of the child during pregnancy. At present, this could only be reliably analyzed by means of invasive prenatal diagnostics (increased risk of miscarriage due to procedures such as amniocentesis or chorionic villus sampling). Against the background of the rapid developments in the field of the analysis of fetal DNA from maternal blood, an analysis of relevant fetal genetic

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characteristics with known influence on placental medication kinetics could perhaps be carried out in the future without increasing  the miscarriage  risk. This would then allow an individually optimal medication selection to be made with the greatest possible safety with regard to fetal exposure.

6.6.2

6

The Sensitivity of the Embryo to Toxic Influences Depends on Its Stage of Development

>>The striking (histo-)morphological and functional changes of the human placenta, especially of the placental barrier, in the course of pregnancy also contribute to the observed temporarily different sensitivity to exogenous influences.

This change is particularly evident when looking at the exposure of the embryo or fetus to antibody molecules in maternal blood. In the first trimester it is assumed that also immunoglobulins of the IgG class (natural “nanoparticles” with a diameter of ~10 nm) do not cross the consistently double-­layered villous trophoblast (7 Chap. 1), but only accumulate in the syncytium. An active physiological transfer of maternal IgG to the fetus only begins in the second trimester. In the third trimester, this transfer develops into a very efficient passive immunization of the fetus to provide for the fetal “nest protection” in the first three months of life (Palmeira et al. 2012). This circumstance has been relevant in fetal-maternal medicine up to now, for example, in connection with the now rare problem of “Rh incompatibility” in Rh-­ negative mothers and Rh-positive fetuses. This can result in varying degrees of fetal hemolysis due to maternal anti-rhesus IgG antibodies (mostly anti-D antibodies). This “morbus  

haemolyticus” develops only after the 18th week of pregnancy, especially if the routine “rhesus prophylaxis” to prevent “immunization” of the mother with fetal erythrocytes was not (successfully) carried out in a previous Rh-positive pregnancy. Otherwise, maternal immunoglobulins still play a pathophysiological role in rare autoimmunological diseases of the pregnant woman with pathological antibody ­production. In systemic lupus erythematosus (SLE), damage to conduction in the fetal heart (atrioventricular block, AV block) by certain maternal Ro/La IgG occasionally occurs from the 16th week of pregnancy onwards, as well as not infrequently to (transient, harmless) neonatal skin manifestations in the sense of “neonatal lupus”. Also in myasthenia gravis, where in the more frequent and harmless case a transient neonatal muscle weakness occurs, severe intrauterine movement disorders of the fetus and consequently permanent limb deformities in the child (“arthrogryposis multiplex congenita”) are sometimes found due to antibodies against the fetal motor endplates. A rare neonatal Graves’ disease, a transient hyperthyroidism caused by autoantibodies (possibly with additional eye problems), has also been described in newborns of affected mothers. In the meantime, an interesting additional aspect has emerged from a medical point of view. Many medications from the group of “biologicals” are recombinantly produced monoclonal antibodies of the IgG class, which are mainly used in oncology, but increasingly also in rheumatic and immunological diseases. The increased use of these very specifically effective medications in women in the fertile phase of life repeatedly leads to unintentional exposure in early pregnancy, especially since experience shows that 30–50% of pregnancies occur unplanned. From a placentological point of view, practically no placental transfer of these therapeutic antibody molecules would be expected in the first trimester and thus in

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the particularly sensitive embryonic development phase. Hence, the teratogenic risk of exposure in early pregnancy would theoretically be classified as very low, unless the antibody causes direct placental damage. However, if the treatment is continued into the second and third trimester, an increasing fetal exposure and, depending on the specific target molecule of the therapeutic antibody, possible adverse effects on the fetus would have to be expected. >>These considerations should also take into account the placenta itself (or its immunological situation) as a possible site of (side) effects of biologicals.

Therefore, in the case of very early exposure to biologicals, the question of a possible increased rate of cardiac malformation arises less than the question of a possible placental pathology in the sense of an increased tendency to miscarriage or an increased risk of FGR (“fetal growth restriction”), preeclampsia or premature birth. From this perspective, a pharmacoepidemiological study from 2009, which, e.g., links the use of TNF-alpha inhibitors on an IgG basis with an increased risk of embryopathies, should be critically  discussed (Carter et  al. 2009). However, it should not go unmentioned here that in analyses of the fluid from the chorionic cavity of early human developmental stages, contrary to the current notion of immunoglobulin transfer in the human placenta, immunoglobulins were also detected, although it remains completely unclear how these can enter this compartment through the early placenta (Gulbis et al. 1992). Ultimately, only the consistent follow-up analysis of a sufficiently large number of children prenatally exposed to biologicals, in comparison to unexposed children of an as adequate as possible comparison cohort, will be able to clarify which human terato-

6

genic risks such biologicals  actually entail, depending on the application during pregnancy (e.g. Weber-Schoendorfer et al. 2015). It would also be desirable that in future these studies take greater account of the possible placenta-associated consequences on prenatal development, which currently receive little attention in  research. Interesting in this context is, for example, the observation of a possible benefit of anti-inflammatory TNF-­ ­ alpha inhibitor therapy in habitual abortions (Winger and Reed 2008). Further evidence of the placental time dependence of teratogenic effects is found in the differential fetal infection rate of transplacentally transmitted infections during pregnancy, depending on the time of infection. This phenomenon can be considered separately from the differences in potential fetal harm from such intrauterine teratogenic infections, depending on their occurrence in relation to embryonic or fetal development. Toxoplasmosis may be mentioned as an example. In the case of a primary infection with Toxoplasma gondii  during pregnancy, an infection of the offspring is observed in ~50% on average, whereby after infection in the first trimester the fetal infection rate is reported to be 15–25%, in the second trimester ~54% and in the third trimester ~65%. Before the 16th week of gestation, infection of the fetus does not seem possible even with maternal initial infection in the first trimester, due to the placental barrier. Nevertheless, the rarer early infection leads to more severe congenital anomalies in the fetus than the later one (Enders 1991). Similarly,  primary cytomegalovirus infections in pregnancy are expected to cause severe damage of the child in ~50% of infected embryos or fetuses when infected in the first trimester, with a transmission rate of ~20%. In the case of maternal infection in the third trimester, placental transmission is found in >80% of cases, but with a significantly lower risk of fetal damage (see RKI Guide for Physicians 2014).

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Interesting in this context is also the dependence of a severe neonatal varicella disease on the existing possibility of “passive vaccination of the fetus” through the placenta. If infection occurs sufficiently long before delivery, anti-varicella IgG produced by the maternal immune system can be transferred across the placenta in efficient amounts, thus preventing severe neonatal disease. The same applies to measles infection and mumps infection during pregnancy, which  only lead to severe neonatal disease when primary infection occurs shortly before birth (Enders 1991). The consequence of teratogenic damage to the placenta itself is in principle also dependent on the time of exposure in the course of pregnancy. Thus, in the case of damage in the first trimester, a reduced vascular remodeling of the spiral arteries with subsequent development of a typical growth restricted, thickened and, at the latest in the third trimester, functionally insufficient FGR placenta (7 Chap. 11) would be possible. Although the same damaging influence in the third trimester would possibly result in a functional disturbance of the placenta or necrosis and thus a supply disturbance of the fetus, the placental morphology would not be impaired in the same way as in the case of early exposure.  

6.6.3

Different Embryotoxic Influences Affect (Embryonic) Development via Relatively Few Specific Mechanisms

>>The most obvious example of the restricted response  variability to exogenous damage is the “all-or-nothing principle” of teratogenic exposure in the 1st and 2nd week of development.

Due to the low specialization of the cells in the early embryo, any  damage can be completely repaired, so that no congenital anomalies are to be expected as a consequence. Alternatively, if the damage is sufficiently severe (not least in the placental tissue, which clearly predominates in terms of quantity in this phase of development!), a clinically inconspicuous early abortion occurs. There is only a normal onset of menstruation at the end of the 2nd week of development (corresponding to the 4th week of pregnancy calculated after the first day of the last menstrual period) at the expected time. In any case, regardless of the nature of the harmful influence (mitotic poison, ionizing radiation in the appropriate dose, infection, etc.), the consequence would be identical. More differentiated, but still limited, are the pathomorphological expressions of the placenta (7 Chaps. 8, 10, 11) in the case of damage from the 3rd week of development until birth. With regard to pathomechanisms of placental damage that have been completely deciphered in molecular detail, it is true, as for pathomechanisms of congenital anomalies in general, that we are far from knowing all of the theoretically limited mechanisms that exist. Often, our current knowledge about exogenously, but also endogenously caused developmental anomalies is based only on empirical data.  

6.6.4

After Exposure to Teratogens, Different Developmental Courses Are Possible in Principle

The fourth basic rule describes the fact that exposure to a potentially teratogenic factor alone does not necessarily lead to congenital health problems. This can be deduced conclusively from the other rules (dependence

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of an effect on individual genetics, the period of exposure, the dose, etc.) and is not least also due to placental phenomena. In addition, however, this rule not only focuses on malformations in the embryo or fetus, but also expands the focus to  additional, more placenta-associated pregnancy problems (miscarriage, prematurity, FGR, etc.). These problems are explicitly considered as a consequence of teratogenic exposure, and the placenta is thus identified not only as a mediator of teratogenic effects, but also as a target organ of such damage. It should be critically noted at this point that research on the use of medication during pregnancy and on the teratogenic effects possibly caused by them regularly examines, in addition to any congenital anomalies, the miscarriage rates and the duration of pregnancy. However, these studies usually do not collect data on the simplest placenta parameters such as placental weight. Likewise, these examinations usually lack ultrasound checks of fetal growth to detect any slowing of intrauterine growth especially in fetuses whose birth weight is not below the 10th percentile, but in whom a bend in the growth curve would indicate FGR and be detectable in this way.

6.6.5

The Way in Which Toxic Influences Reach the Embryo/Fetus Depends on Their Physical and Chemical Properties

This rule refers directly to pharmacokinetics at the placental barrier. Only a few teratogenic influences, such as ionizing radiation, whose negative influence also depends on the physical properties of the radiation (alpha, beta, gamma rays, other particle rays, ionization density, wavelength, dose), or possible ascending infections from the vagina, which can also reach the amniotic fluid directly via the fetal membranes,

6

bypassing the placenta, are not influenced by the physicochemical interaction with the placental barrier in their effect on embryonic or fetal development. Little is known about the efficiency of substance transfer in the early human placenta or in the second trimester. The ­situation of the mature placenta in the third trimester, on the other hand, can be simulated comparatively well in this respect (7 Chap. 15) or determined in  vivo (by comparing the maternal blood level with the fetal umbilical cord blood level measured immediately after birth).  

>>In general, a molecular mass 17,000 D), should be expected to have a comparatively very low infant “co-­ treatment”. Likewise, fibrinolysis by means of alteplase (molecular mass of >59,000 D) should be limited mainly to the maternal circulation. Active placental transport mechanisms, which can ensure that large hydrophilic molecules with a mass of 150,000 D are efficiently transported from the maternal blood into the fetal circulation, play an important role for the placental barrier. This can be seen in the example of IgG transport in the

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second and third trimesters, where the IgG Fc part is specifically channeled through the syncytium via a neonatal Fc receptor (FcRn) and receptor-mediated transcytosis (point 2 “The sensitivity of the embryo to toxic influences depends on its developmental stage”).

gen production in the adrenal gland of female fetuses and thus to prevent virilization of the external genitalia in cases of prenatally diagnosed adrenogenital syndrome (AGS).

>>Besides specific transport mechanisms for very large molecules, placental metabolism as well as transport proteins from the group of multi-drug resistance proteins have to be considered.

6.6.6

Multi-drug resistance proteins (specifically BCRP, “breast cancer resistance protein”) ensure that e.g. the blood sugar-lowering substance glibenclamide is not transferred in significant quantities across the placenta despite good oral bioavailability and small molecular weight, in contrast to other similar active substances from the sulfonylurea group. The agent is exported quite efficiently by active transport, from the trophoblast into the maternal circulation via BCRP molecules. In addition, high plasma protein binding and a short half-life of glibenclamide in the blood also contribute to the fact that the drug does not transfer appreciably into the fetal circulation and is discussed as a possible alternative to insulin therapy in type 2 diabetes or gestational diabetes in pregnancy. The difference in placental inactivation between halogenated glucocorticoids such as betamethasone and dexamethasone on the one hand and non-halogenated ones such as cortisol or prednisolone on the other hand is responsible for the fact that only about 10% of a maternal prednisolone concentration is measured in the umbilical cord blood, but about 30% of a beta- and almost 100% of a dexamethasone concentration (Schäfer et  al. 2012). This finding can be used to prioritize prednisolone when maternal therapy is needed during pregnancy. In contrast, dexamethasone would be the medication of choice for infant glucocorticoid requirements, e.g. to reduce increased andro-

Dose-response Relationships Apply in Teratology as Elsewhere in Pharmacology and Toxicology

The old basic rule of toxicology “the dose makes the poison” is of course also valid for placental as well as for embryonic or fetal exogenously caused damage. Any exposure, regardless of the substance, can be toxic and thus teratogenic, or non-toxic, depending on the dose. . Figure 6.2 illustrates this basic principle. The placenta usually plays a key role for the embryonic or fetal dose, in addition to the fetal liver metabolism and a possible substance accumulation in the amniotic fluid. As previously discussed in the context of the other Wilsonʼs rules, the fetal dose does not depend exclusively on the maternal blood level.  

>>Placental genetics as well as developmental placental structure and physiology and the particular species-specific placental barrier function significantly influence the fetal dose and thus ultimately a teratogenic effect.

In the case of a teratogenic effect, a threshold dose (NOAEL, “no observable adverse effect level”) must be assumed, above which teratogenic effects become apparent, depending on the genetically determined individual sensitivity. Below this “minimum dose”, the risk of congenital anomalies is not increased. Unfortunately, NOAEL values for most substance exposures are still unknown. This is also due to the fact that in

6

119 Teratology

?

Therapeutic range

? 100 %

Non-toxic range

Teratogenic range

Lethal range for embryo/fetus

Toxic range for mother/dam

Basic risk

Damaged embryos/dams

0% variable, species- specific or individual threshold dose range

Increasing dose

..      Fig. 6.2  Dose-response relationships in teratology determined by animal experiments. (Modified after Wilson 1977 and Schäfer et al. 2012)

most observational studies of medication exposure during pregnancy, although the amount of medication taken is recorded, maternal drug blood levels are not determined. Hence, no clear correlation can be established between teratogenic effects and drug dose. This would only be possible if the individual maternal pharmacokinetics were taken into account, as well as the often noticeably reduced tendency during pregnancy to actually take the prescribed amount of medication (Lupattelli et  al. 2014). Both could be easily objectified on the basis of measured medication levels in the blood, to effectively document at least the placental substance load. For a few drugs that show a significantly altered metabolism and excretion due to pregnancy, the recommendation to regularly monitor serum levels during pregnancy in the sense of “therapeutic drug monitoring” (TDM) has become established to avoid under- or overdosing. Therapy with the antiepileptic drug lamotrigine or treatment with lithium would be examples of this (Schäfer et  al. 2012). Currently, a new dimension is emerging in teratology. The question is increasingly being asked as to which prenatal influences affect the long-term health of the child in

later life, i.e. the tendency to high blood pressure, obesity, diabetes, arteriosclerosis or cancer, psychosocial and intelligence development, the predisposition to psychiatric diseases, immunological development and the ageing process. A theoretical basis for these considerations is provided by the Barker or DOHaD (Developmental Origins of Health and Disease) hypothesis (Wadhwa et al. 2009). This hypothesis initially drew attention to the possible influence of factors during prenatal development on later disease risks, based on epidemiological observations of associations between maternal nutritional problems in pregnancy and the risk of cardiovascular disease in children decades later. Epigenetic changes are under discussion as important pathomechanisms  responsible for these observations, which, given a genetic background, allow for sustained exogenously induced modulability, so that prenatally induced long-term functional maladaptations and disorders appear possible. Such epigenetically mediated phenomena could also be caused as early as the all-or-nothing phase, as discussed in connection with teratogenic effects and epigenetic changes caused by in vitro fertilization (IVF) methods (Wilkins-Haug 2009).

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Conclusion

the American association for the advancement of Science, on 29. Dec, Boston/MA) Lupattelli A, Spigset O, Nordeng H (2014) Adherence In summary, it can be stated that in the old to medication for chronic disorders during pregteratological question of the causes of connancy: results from a multinational study. Int J Clin Pharm 36(1):145–153 genital health problems in a variety of ways, Palister JL, Paré A (1982) On monsters and marvels. the placenta plays a relevant role in answerUniversity of Chicago Press, Chicago ing it. This multifaceted involvement of the Palmeira P, Quinello C, Silveira-Lessa AL, Augusta placenta in teratology is in part already Zago C, Carneiro-Sampaio M (2012) IgG placenquite clearly definable, and an understandtal transfer in healthy and pathological pregnancies. Clin Dev Immunol 2012:985646 ing of its importance is now medically appliPhelan MC, Pellock JM, Nance WE (1982) Discordant cable. In many areas, however, there is expression of fetal hydantoin syndrome in heteroobviously still a great need for research, and paternal dizygotic twins. N Engl J Med 307(2):99– we can certainly expect further diagnosti101 cally or therapeutically useful placentologi- RKI-Ratgeber für Ärzte (2014) Zytomegalievirus-­ Infektion, Stand 20.01.2014. https://www.­rki.­de/ cal findings in teratology in the future. DE/Content/Infekt/EpidBull/Merkblaetter/ Ratgeber_Zytomegalievirus.­html. Accessed on 10 Nov 2017 References Rösch C, Steinbicker V (2003) Aetiology of congenital malformation, analysis of malformation registry data compared with the Kalter and Warkany Bliek BJ, van Schaik RH, van der Heiden IP, Sayed-­ study (abstract). Reprod Toxicol 17:503 Tabatabaei FA, van Duijn CM, Steegers EA, Steegers-Theunissen RP, Eurocran Gene-­Schäfer C, Spielmann H, Vetter K, Weber-­Schöndorfer C (2012) Arzneimittel in Schwangerschaft und Environment Interaction Group (2009) Maternal Stillzeit, 8. Aufl. Urban & Fischer, Elsevier, medication use, carriership of the ABCB1 3435C München, S 5 > T polymorphism and the risk of a child with cleft lip with or without cleft palate. Am J Med Schardein JL (2000) Chemically induced birth defects, 3. Aufl. Marcel Dekker, New York Genet A 149A(10):2088–2092 Carter JD, Ladhani A, Ricca LR, Valeriano J, Vasey Schumacher GH, Fanghänel J, Persaud TV (1992) Teratologie 1992, Kapitel 2 Geschichte der FB (2009) A safety assessment of tumor necrosis Teratologie, S 18–49 factor antagonists during pregnancy: a review of the Food and Drug Administration database. J Wadhwa PD, Buss C, Entringer S, Swanson JM (2009) Developmental origins of health and disease: Rheumatol 36(3):635–641 brief history of the approach and current focus on Elwyn G, Frosch D, Thomson R, Joseph-Williams N, epigenetic mechanisms. Semin Reprod Med Lloyd A, Kinnersley P, Cording E, Tomson D, 27(5):358–368 Dodd C, Rollnick S, Edwards A, Barry M (2012) Shared decision making: a model for clinical prac- Weber-Schoendorfer C, Oppermann M, Wacker E et  al (2015) Pregnancy outcome after TNF-α tice. J Gen Intern Med 27(10):1361–1367 inhibitor therapy during the first trimester: a proEnders G (1991) Infektionen und Impfungen in der spective multicentre cohort study. Br J Clin Schwangerschaft, 2. Aufl. Urban & Pharmacol 80(4):727–739 Schwarzenberg, München Friedman JM (2010) The principles of teratology: are Wilkins-Haug L (2009) Epigenetics and assisted reproduction. Curr Opin Obstet Gynecol they still true? Birth Defects Res A Clin Mol 21(3):201–206 Teratol 88(10):766–768 Gulbis B, Jauniaux E, Jurkovic D, Thiry P, Campbell Wilson JD (1977) Embryotoxicity of drugs in man. In: Wilson JD, Frazer FC (Hrsg) Handbook of S, Ooms HA (1992) Determination of protein patTeratology, Bd 1. Plenum Press, New York, S 309– tern in embryonic cavities of human early preg355 nancies: a means to understand materno-embryonic exchanges. Hum Reprod Winger EE, Reed JL (2008) Treatment with tumor necrosis factor inhibitors and intravenous immu7(6):886–889 noglobulin improves live birth rates in women Lorenz EN (1972) Predictability: does the flap of a with recurrent spontaneous abortion. Am J butterflyʼs wings in Brazil set off a tornado in Reprod Immunol 60(1):8–16 Texas? (Address at the 139th Annual Meeting of

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The Effects of Legal and Illegal Drugs on Placental Function Justine Fitzgerald and Ekkehard Schleußner

Contents 7.1

Introduction – 122

7.2

Smoking During Pregnancy – 122

7.2.1 7.2.2 7.2.3 7.2.4 7.2.5

T obacco Ingredients – 123 Effects on Placental Morphology – 123 Effects on Trophoblast Cells – 125 Oxidative Stress and Endothelial Dysfunction – 125 Placental Transcriptome – 126

7.3

Alcohol During Pregnancy – 126

7.3.1 7.3.2 7.3.3

E ffects on Placental Morphology – 127 Effects on Trophoblast Cells – 128 Oxidative Stress and Endothelial Dysfunction – 128

7.4

Methamphetamines and MDMA – 128

7.5

Cocaine – 130

7.6

Opiates – 130

7.6.1 7.6.2

 lacental Transfer – 131 P Effects on Trophoblast Cells – 131

7.7

Cannabis – 131

7.7.1 7.7.2

 lacental Transfer – 132 P Effects on Trophoblast Cells – 132

References – 133 © Springer-Verlag GmbH Germany, part of Springer Nature 2023 B. Huppertz, E. Schleußner (eds.), The Placenta, https://doi.org/10.1007/978-3-662-66256-4_7

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7.1

7

J. Fitzgerald and E. Schleußner

Introduction

According to the Ministry of Health, 1,800,000 people in Germany were dependent on alcohol in 2015, and an estimated 2,300,000 people were dependent on medication. Around 600,000 people display a problematic use of cannabis and other illegal drugs. Most recently, data were published in February 2017, according to which 2,650,000 children in Germany live in families with addiction problems, and of these, 60,000 live in families with illegal drug use, although it is assumed that the number of unreported cases is significantly higher (Moesgen et al. 2017). For most pregnant women, pregnancy is a special phase of life in which they responsibly decide to do everything possible to ensure the optimal development of the unborn child, in particular by stopping smoking and drinking alcohol or abstaining from drug use. For the vast majority, any drug use is obsolete, but for an unfortunately not diminishing minority, this is not the case. Reliable German data do not exist, for the USA a constant figure of 5.9% pregnant women with illegal drug use is reported (Ross et al. 2015). Prenatal drug exposure poses a variety of risks to mother and child that can lead to obstetric and neonatal complications. The most common problems arise due to disturbances in placental function: fetal growth restriction (FGR), preeclampsia, prematurity, premature rupture of membranes (PROM) to premature placental abruption and intrauterine fetal death (IUFD). The placenta-associated effects of the major licit and illicit drugs and possible pathomechanisms in the placenta are described below.

7.2

Smoking During Pregnancy

The WHO estimates that smoking during pregnancy is the most important preventable risk factor for a number of pregnancy complications. While FGR, preterm birth and stillbirth are significantly more common, preeclampsia occurs less frequently in pregnant women who smoke. Although overall tobacco use, particularly during pregnancy, has declined in most developed countries in recent decades, prevalence is still thought to be 10–20%. Pregnant women who smoke are often younger, single or have a partner who also smokes, have unplanned pregnancies, are not employed and have a lower socioeconomic and educational status. Interestingly, smoking is less prevalent among women of reproductive age with a migration background (German Ministry of Health [Bundesministerium für Gesundheit] 2016). Through targeted intervention at the beginning of pregnancy, it is realistic to reduce the number of smokers to about half. The most promising results were achieved in studies that offered counselling and problem-­solving strategies and in those that used incentive measures (e.g. vouchers for special additional services) and personal feedback (ultrasound monitoring of normal child development, determination of cotinine levels in urine as a nicotine breakdown product and thus direct evidence of nicotine consumption). In its recommendations, the German Medical Association has adopted the so-called 5A scheme of the WHO (Ask-­Advise-­Assess-Assist-Arrange), according to which tobacco consumption can and should be addressed at the beginning of pregnancy. Non-individualized health campaigns tended to be less successful. Meta-­ analyses also demonstrate that

123 The Effects of Legal and Illegal Drugs on Placental Function

about half of those women who temporarily abstained from tobacco smoking during pregnancy resume smoking within the first 6 weeks to 6 months after pregnancy. 7.2.1

Tobacco Ingredients

Tobacco consists of a mixture of chemical substances, of which for nicotine, carbon monoxide CO, cadmium and benzopyrene the effect on the placenta have been studied. Nicotine crosses the placental barrier unhindered and, in the case of chronic abuse, reaches the same levels in the fetal blood as in the mother (Lambers and Clark 1996). Nicotinic receptors are found in almost all placental cells, including the syncytiotrophoblast, the Hofbauer cells, the endothelium and the vascular musculature (Lips et al. 2005); hence, it can be assumed that nicotine can influence placental vascularisation and perfusion as well as nutrient transfer. Cotinine passes just as readily into the fetal circulation and can accumulate there at nearly twofold concentrations, as placental perfusion studies have shown (Sastry 1991). Cadmium acts as a metalloestrogen and endocrine disruptor in reproductive organs and during fetal development (Kawai et al. 2002). It accumulates in the placenta, resulting in twice as high tissue levels in smokers (Stasenko et  al. 2010). It impairs placental hormone synthesis of both steroid and proteohormones. Benzopyrenes are polycyclic aromatic hydrocarbons which, in high concentrations in cigarette smoke, can exert mutagenic effects in both active and passive smoking

7

(Lee et al. 2011). They also pass via the placenta into the fetal blood and apparently modify placental metabolism (Sanyal and Li 2007). Carbon monoxide (CO) is found in elevated blood concentrations in smokers, readily crosses the placental barrier and, like nicotine, can reach high fetal levels in the fetal compartment. It can ultimately cause placental and fetal hypoxia through the formation of carboxyhemoglobin (Rogers 2009). The effects of tobacco consumption on the pregnant woman, placental morphology and function, and the fetus are shown schematically in . Fig. 7.1.  

7.2.2

Effects on Placental Morphology

Regardless of the type of tobacco consumption, smokers have smaller placentas with thicker villous membranes and increased trophoblast volume. The outer syncytiotrophoblast shows degenerative changes with increased apoptosis, necrosis and syncytial knots. The intervillous space is relatively reduced and at the same time, the villous capillaries are shortened with a smaller exchange surface (Ashfaq et al. 2008). These micromorphological changes reduce the transport capacity for nutrients and gas exchange. Placental effects can be detected directly by Doppler sonography of maternofetoplacental perfusion. Dose-dependently, uteroplacental perfusion is worsened, umbilical artery resistance is increased and ultimately birth weight is decreased (de Machado et al. 2011) (. Fig. 7.2).  

Uteroplacental perfusion - Doppler: transiently elevated uteroplacental RI - Altered vasoconstriction of the uterine arteries - Endothelial dysfunction of the uterine endothelial cells - Reactive vasorelaxation due to higher endogenous CO levels at the placental basal plate

Fetoplacental perfusion - Doppler: increased resistance in the umbilical arteries (RI) - Higher vasoconstriction of fetal vessels - Lower tolerance to oxidative stress

Endocervical immunity - Increase in anti-inflammatory cytokines in the 1st trimester - Cytokine profile: not changed despite vaginosis - Langerhans cells: Decreased in number - Lymphocytes: no effects in non-pregnant women

Placental immunity - Inhibition of proinflammatory cytokines - Increased number of NK-cells in induced abortions in a mouse model - Macrophages, possibly, increased, - Inhibition of an enzyme that breaks down labor-stimulating factor

..      Fig. 7.1  Possible effects of smoking during pregnancy on placenta-associated compartments

Placental villous development - Lower placental weight with unchanged placental volume - Increased trophoblast volume - Relative reduction of the intervillous space of the peripheral villous mass and its surface area - Increased aging & degeneration: ο Syncytiotrophoblast thinning and necrosis ο Increased syncytial knots ο Increased collagen in anchoring villi ο Cellular alterations in the syncytiotrophoblast - Cytotrophoblastic stem cell layer & villous membrane thickened - Vasculosyncytial membranes thinner - Shorter length of fetal capillaries and reduced capillary surface, thus smaller exchange area

Maternal immune system - Increased neutrophils, monocytes & lymphocytes - Between 14 and 20 weeks of gestation: ο elevated CD3+ T lymphocytes ο increased CD4+ T lymphocytes ο CD8+ T lymphocytes unchanged ο Drop in CD56+ NK cells ο Later normalization of all changes - Decrease of adhesion molecules for monocytes and granulocytes

Trophoblast cells - Apoptosis: inconsistent data - Reduced number of trophoblast cell columns starting from anchoring villi reach the decidua - Lower number of cytotrophoblast columns of the adhesive villi reach the decidua - Inhibition of differentiation into invasive trophoblasts - Reduced capacity for invasion/migration - Reduced synthesis of hCG, HPL, estriol and estradiol - Increased PIGF secretion - Altered metabolism of placental peptide hormones - Reduced cellular energy synthesis - Enhanced adaptation to oxidative stress

7

Maternal vascular system - Elevated pulse and blood pressure - Amplif ied inf lammation markers - Increased markers of endothelial dysfunction - Inhibited IL-6 expression in endothelial cells.

124 J. Fitzgerald and E. Schleußner

125 The Effects of Legal and Illegal Drugs on Placental Function

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80 70 Percentile (mean)

60 50 40 30 20 10 0 medium RI AU

RI UA

RI MCA

Birth weight

Carbon monoxide in the exhaled air 0 ppm

1–4 ppm

..      Fig. 7.2  Dependence of the placental perfusion resistance (resistance index, RI) on the maternal side (uterine artery, AU), the fetal side (umbilical artery, UA), the fetal cerebrovascular resistance (middle cerebral artery, MCA) as well as birth weight on the car-

7.2.3

Effects on Trophoblast Cells

Smoking may have different effects on the apoptosis rate. While increased trophoblast apoptosis was demonstrated in smokers, lower apoptosis was found in the syncytiotrophoblast. Carbon monoxide, on the other hand, has anti-apoptotic and anti-necrotic effects, at least in an in  vitro model (Bainbridge et al. 2005). Smoking appears to affect the balance between proliferation and differentiation of trophoblasts and particularly impairs the differentiation of invasive cytotrophoblasts (Jauniaux and Burton 2007). Cadmium decreases trophoblast proliferation, and benzopyrenes negatively affect trophoblast stem cell transcription (Xie et al. 2010). However, not only differentiation into an invasive phenotype, but also trophoblast invasion directly is  inhibited. Instead of invading the decidua starting from anchoring villi, a larger proportion of anchoring villi fail to reach the uterus and degenerate in the intervillous space (Genbacev et  al. 2000). Nicotine inhibits I-selectin-mediated

5–9 ppm

>10 ppm

bon monoxide concentration of exhaled air as a measure of smoking. The CO content of exhaled air is measured in parts per million (“ppm”). (Modified after de Machado et al. 2011) (Graphical representation of numbers, column chart)

adhesion as well as the expression of fibronectin and its receptor factors, which are required for undisturbed migration and invasion of extravillous trophoblasts. Placental hormone production of hCG (human chorionic gonadotropin), hPL (human placental lactogen), estriol and estradiol is lower in smokers, while the angiogenesis factor PlGF (“placental growth factor”) is found increased in maternal serum (Zhang et al. 2011). 7.2.4

Oxidative Stress and Endothelial Dysfunction

The various tobacco components cause oxidative stress in the placenta. Maternal cotinine levels correlate with markers of maternal oxidative stress as well as with markers of placental DNA oxidation. The generation of free radicals simultaneously activates the antioxidant system. In vivo studies demonstrate increased expression of heme oxygenase enzymes (HO-1 and HO-2) in the placental basal

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plate of smokers, while in vitro HO-1 expression has been shown to be dose-dependent in trophoblasts (Sidle et  al. 2007). HO-1 shows anti-inflammatory properties in addition to its antioxidant properties (Tranquilli and Landi 2010). HO expression is also increased several times in the myometrium of pregnant women, where it reduces both spontaneous and oxytocin-induced contractility (Acevedo and Ahmed 1998). Nicotine reduced sFlt-1, s-endoglin and PlGF release in trophoblast cultures, whereas this was increased in HUVEC (Human Umbilical Vein Endothelial Cells) cultures (Romani et al. 2011). CO decreased sFlt-1 and s-endoglin production in endothelial cells and placental villous explants from preeclamptic pregnant women (Cudmore et al. 2007). Furthermore, the activity of endothelial nitric oxide synthase (eNOS) in placental villous tissue and the umbilical endothelium is reduced in pregnant women who smoke, which may lead to a disturbance of NO-­ dependent vascular dilatation and thus directly to fetal insufficiency (Myatt et  al. 1997). In the pathogenesis of preeclampsia, an angiogenic imbalance between growth factors such as VEGF (“vascular endothelial growth factor”) and PlGF and their vascular wall receptors (Flt-1) or soluble receptors (sFlt-1, s-endoglin) plays a central role. The concentration of the latter is substantially regulated by placental HO-1, as shown above. In addition, a higher concentration of PlGF has been demonstrated in female smokers. Via these mechanisms, the sFlt-­1/PlGF ratio  is reduced. This might explain why smokers selectively have a lower incidence of that form of preeclampsia, which is not associated with placental insufficiency and intrauterine growth restriction. However, the percentage of cases of early preeclampsia (35 years, pelvic irradiation, reproductive interventions such as in vitro fertilization, and female gender of the fetus are associated with an increased risk of developing PAS (Henrich et al. 2008; Doumouchtsis and Arulkumaran 2010; Esh-­Broder et al. 2011; Fitzpatrick et al. 2012; Hayashi et al. 2012; Nageotte 2014; Timor-­ Tritsch et  al. 2014; Kaser et  al. 2015; Silver et  al. 2015). The most common constellation of PAS  is placenta praevia following cesarean delivery (Wu et al. 2005) (. Table 8.1).  

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.       Table 8.1  Risk factors and associated incidence of the placenta accreta spectrum (PAS). (Clark et al. 1985; Miller et al. 1997; Silver et al. 2006a, b; NIH 2010) Risk factor

Risk of PAS (%)

One previous C-section

0.3

Two previous C-sections

0.6

Three previous C-sections

2.4

Placenta praevia and

8

– no previous C-section

1–5

– one previous C-section

11–25

– two previous C-sections

35–47

– three previous C-sections

40

– four or more previous C-sections

50–67

8.1.4

Definition

In defining the implantation disorder of the placenta, a distinction is made between the histopathological definition obtained postpartum and the prepartum and peripartum clinical definition. Histopathologically, a differentiation can be made between placenta accreta, increta and percreta depending on the extent and depth of invasion of the chorion frondosum into the myometrium (. Fig. 8.1). Placenta accreta shows invasion of the trophoblast to the myometrial inner wall with disruption or absence of the decidua basalis. Placenta increta shows invasion of the trophoblast deep into the myometrium but not beyond the serosa, and placenta percreta shows invasion up to the uterine serosa or beyond the uterine borders and the serosa with or without infiltration into neighboring organs such as the urinary bladder, the parametria, or the intestine. In an analysis of a total of 138 histologically confirmed PAS cases, the frequency distribution was as follows: placenta accreta 79%,  

..      Fig. 8.1  Schematic representation of different degrees of severity (depth of invasion, localization and vascular extent) of the placenta accreta spectrum. (1) Placenta accreta with invasion of the trophoblast beyond the decidua basalis to the border of the myometrium; (2) Placenta increta with invasion of the trophoblast deep into the myometrium; (3) Placenta percreta with invasion of the trophoblast into the myometrium to the border of the serosa or beyond with or without infiltration of adjacent structures; (4) Placenta increta/percreta with cervical invasion and bladder wall involvement

placenta increta 14% and placenta percreta 7% (Miller et al. 1997; Wu et al. 2005). According to the clinical relevance, however, only two groups are distinguished, placenta accreta and placenta increta/percreta. In the case of placenta accreta, which requires a manual detachment or curettage, the placenta can be removed completely and no further surgical interventions are necessary. In the case of placenta increta/percreta, the placenta cannot be completely removed by manual detachment or post curettage. Further surgical interventions are required (Henrich and Braun 2013), and these are associated with increased maternal mortality of up to 7% overall (O’Brien et al. 1996). This must be distinguished from a placenta which, in the area of scar dehiscence, for example in the area of the old C-section scar, bulges out onto the surface of the uterus and is only covered by the uterine serosa (. Fig. 8.2). This is not a classical PAS and its treatment is usually much less complicated.  

139 Placenta-Related Hemorrhage: Pathophysiology, Diagnostics, Management

a

8

b

Dehiscence Myometrium

UB

..      Fig. 8.2  a, b Uterine scar dehiscence. a Transabdominal sonography. b Intraoperative findings. UB Urinary bladder

>>In the English literature, all degrees of severity are often summarized under the generic term “placenta accreta”. This complicates the comparability of studies with clearly different clinical significance.

8.1.5

Pathogenesis

The exact pathogenesis of PAS  is not yet known. One theory describes a very thin, interrupted or missing decidua basalis as the cause (Khong 2008) and attributes this to previous intrauterine interventions (Tantbirojn et al. 2008). After all, one study showed an association with previous intrauterine interventions such as cesarean section, curettage and/or myoma enucleation in 80% of PAS  cases (Tantbirojn et  al. 2008). Other theories assume an overshooting, non-­regulated extravillous trophoblast invasion or a disturbed maternal vascular remodeling in the area of a uterine scar (Wehrum et al. 2011; Chantraine et al. 2012; Jauniaux and Jurkovic 2012). The duration of pregnancy seems to play a minor role in this regard for the depth of implantation, as there are already histopathologically confirmed findings of a placenta percreta from

the 14th week of gestation (Pron et al. 2005; Hamar et al. 2006). Another theory for the development of PAS is based on the preferential nidation of the placenta in the area of hypoxemic tissue areas, which are found in particular in scar tissue after intrauterine surgery (Rosen 2008). 8.1.6

Diagnosis

The antepartum diagnosis of PAS remains a challenge for the investigator even today (Tutschek et al. 2014). In a large population-­ based study, PAS was not detected ­antepartum in a total of 50–70% of cases (Tikkanen et al. 2011; Thurn et  al. 2016). However, it is the prenatal diagnosis of PAS and the interdisciplinary birth planning based on this diagnosis that allows the significantly improved surgical outcome and maternal survival. For example, antepartum diagnosis decreases maternal blood loss compared to pregnant women in whom PAS  was only discovered perioperatively (Warshak et  al. 2010; Tikkanen et  al. 2011; Chantraine et  al. 2013). The antepartum differentiation of the severity of PAS, in particular the differentiation of placenta accreta versus placenta increta with the

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resulting clinical consequences, remains difficult despite increasing awareness of this issue. In addition to taking a detailed medical history and recording the known risk factors (. Table  8.1), prenatal ultrasound and, in addition, MRI are used for diagnostic purposes. Ultrasound diagnosis of PAS by means of B-scan and color or power Doppler sonography offers an irreplaceable imaging procedure to detect this life-threatening disease. In general, it is recommended to perform transabdominal and transvaginal sonography when the urinary bladder is moderately filled. The use of 12 MHZ transducers allows a high resolution especially in the near field. Already in early pregnancy between 5 and 7 weeks of gestation, a scar pregnancy should be ruled out and the position of the chorion in relation to the sectional scar should be assessed sonographically (. Fig. 8.3) (Armbrust et al. 2015). A scar pregnancy in the first trimester with either an anterior myometrial thickness 3  cm (7 Sects. 8.1.6 and 9.1). significantly reduces the likelihood of vaginal bleeding and/or emergency cesarClinical Management ean section (Stafford et al. 2010; SekiguThe following pragmatic approach applies chi et al. 2015). to asymptomatic women without bleeding 5 5 Thickness of the placental margin: The (Reddy et al. 2014): thickness of the placental margin over The placental location should be considthe internal cervical os (thickness of >1 ered normal if the placental margin is >2 cm cm) also appears to be a good predictor from the internal cervical os in pregnancies of vaginal bleeding (Ghourab 2001; from 16 + 0 weeks gestation. If the placental Saitoh et al. 2002). margin is 3 cm and placental margin < 1 cm outpatient management, C-section from 37+0 weeks gestation • Cervical length 1cm Pulmonary maturation induction and closer monitoring • Cervical length 2 cm

Distance 1–2 cm

Distance 1 cm

Vaginal delivery at term

Follow-up checks every 1-2 weeks

Therapy like placenta praevia

Cervical length 1 cm close monitoring and C-section from 37+0 weeks gestation Cervical length >25 mm and placental margin 35 years

+

FGR fetal growth restriction; SGA small for gestational age; TTTS twin-to-twin transfusion syndrome (According to Ghaheh et al. 2013; Gelaye et al. 2016; Lanna et  al. 2017; Cheng et  al. 2012; Mendola et  al. 2013; Aliyu et  al. 2011; Gul et al. 2016; Khattak et al. 2012; Markhus et al. 2011; Minna et al. 2011; Pariente et al. 2011)

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tact bleeding or bleeding in the context of a uterine rupture. If a diagnosis of premature placental abruption is made, the risk for preterm delivery increases four to six times. The risk for stillbirth increases 8.9- to 12-fold (Ananth et  al. 1999; Ananth and Wilcox 2001). The risk of maternal death peripartum increases sevenfold with premature placental abruption compared with the overall maternal mortality rate. 8.4.2

8

Definition

Premature placental abruption describes a detachment of the placenta before delivery of the child from the 21st week of pregnancy (SSW) onwards. A retroplacental hematoma of varying size or extent forms in the decidual layer. In the course of the detachment process, a partial placental abruption may develop into a complete abruption with considerable consequences for mother and child. The clinical spectrum and the consequences for mother and child are correspondingly large. Detachment of more than half of the placenta is often associated with intrauterine fetal death (IUFD) (Ananth et  al. 1999). Loss coagulopathy forms as the bleeding progresses, which is then complicated by disseminated intravascular coagulopathy (DIC). In the context of coagulopathies, there is an unrestrained consumption of coagulation factors, ­opening the door to further complications (atony, uterine bleeding, need for hysterectomy, etc.). 8.4.3

Etiology

Immunological, ischemic, and/or inflammatory stimuli are thought to be responsible for the onset of premature placental abruption, leading to rupture of decidual arteries in the basal layer, resulting in the buildup of the hematoma between the uterine wall and placenta that is typical of placental abrup-

tion (Ananth et  al. 2006a; Matsuda et  al. 2011; Tikkanen 2010). The leaking blood spreads into the decidual layer and dislodges the placenta from its adhesion with the uterus (dissection). Depending on how far this process progresses, the exchange of gases and nutrients between mother and fetus  – i.e. placental function  – is permanently disrupted. When a critical exchange area is reached, the lost placental function can no longer be compensated for, and the fetus is undersupplied. In addition, the progressive blood loss endangers the health of the mother. The development of premature placental abruption can have various causes. Trauma (abdominal trauma, fall, minor trauma, etc.) leads to shear forces between the placenta and the uterine wall, which can cause premature placental abruption (Ananth et al. 2006a, b). The risk of premature placental abruption is increased even with minor trauma (Cheng et  al. 2012). On the other hand, uterine scarring (e.g., condition following myoma enucleation) and/or synechiae or myoma may contribute to insufficient formation of the decidual layer in some circumstances, thus promoting premature placental abruption. Why cocaine increases the incidence of premature placental abruption is unknown. Vascular constriction leading to acute circulatory disturbances in the decidua is discussed here, with subsequent ischemia, necrosis, and hemorrhage (Mbah et  al. 2012). Last but not least, this process is also conceivable in cases of excessive elevations of maternal blood pressure or in cases of chronic placental insufficiency with significantly impaired placental perfusion. Thus, pregnant women with hypertensive disease have a fivefold increased risk of severe premature placental abruption compared with normotensive pregnant women. This perfusion disturbance can also occur in nicotine abuse during pregnancy and thus lead to ischemia in the decidual layer, with the consequences of

165 Placenta-Related Hemorrhage: Pathophysiology, Diagnostics, Management

premature placental abruption (Kaminsky et  al. 2007). Interestingly, recent data suggest that administration of vitamins C and E may reduce the rate of premature placental abruption and preterm birth in nicotine abusers in pregnancy in the normal collective (Abramovici et al. 2015). The extent to which thrombophilia (MTHFR gene mutation, hyperhomocysteinemia, factor V Leiden mutation) increases the risk of premature placental abruption is a matter of considerable controversy in the literature (Tikkanen 2011). Except for the development via trauma, premature placental abruption must ultimately always be interpreted as a process in connection with a chronic change in the placenta, which leads to a change in the vessels in the decidual layer, resulting in ischemia and inflammatory reactions and thus in the development of necroses, infarcts and ultimately hemorrhages. Thrombin plays a key role in the development of premature placental abruption. Thrombin can contribute directly or indirectly to uterine activation of the myometrium (contractions) (Elovitz et al. 2000; Lockwood et al. 2012). In addition, it interferes with the coagulation cascade in the mother and promotes the development of disseminated intravascular coagulopathy (DIC) (Thachil and Toh 2009).

8.4.4

Clinical Signs

There are neither typical clinical signs nor a typical regular course for premature abruption of the placenta. It can be subacute or chronic. In about 80% of cases it is associated with vaginal bleeding. In two thirds of the cases, a hard uterus (wooden uterus) is present. CTG changes are documented in 60% of cases (Hurd et al. 1983). In 20–30% of all cases, the pregnant woman shows no symptoms at all (van de Vondel 2010). A typical symptom is abdominal pain, but also back pain, especially if a posterior placenta

8

is present. In addition, a sudden drop in blood pressure in the pregnant woman can be indicative. Mostly, the clinical picture is diffuse, which leads to a delayed diagnosis and initiation of further steps. This is especially the case if the lower abdominal or back pain is not associated with visible vaginal bleeding (Suzuki 2015). It is not uncommon for premature placental abruption to be diagnosed at the time of delivery (e.g. during cesarean section). However, the signposts for diagnosis are usually lower abdominal pain and vaginal bleeding. The severity of the bleeding does not correlate with the degree of detachment of the placenta and/ or fetal vulnerability (Kasai et  al. 2015). There is evidence that lower abdominal pain, as a symptom of incipient placental abruption, may underlie circadian rhythms with a higher rate of intrauterine fetal death in the morning hours (Ohhashi et al. 2017). Therefore, such complications should be expected especially during periods of lower staffing density in clinics. In addition to the typical clinical signs, careful attention should be paid to risk factors  during diagnosis (. Table  8.6) and preterm labor that cannot be explained in any other way (possibly hyperfrequency). For example, if you are caring for a patient with severe preeclampsia, you must expect such a complication at any time and correctly interpret the signs – as opposed to the prodromes typical of preeclampsia.  

8.4.5

Instrumental Diagnostics

B-Mode Sonography Although B-mode ultrasonography usually provides a good view of the placenta, the diagnosis of premature placental abruption is not easy. The expected sonographic image of an echo-poor space between the inner wall of the uterus and the placenta does not show up regularly. The developing hematoma can be hypo-, hyper- or isoechogenic

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..      Fig. 8.24  Premature partial placental abruption. The retroplacental hematoma with a diameter of 5 cm was measured in the 32nd week of gestation. The hematoma is located in the periphery of the placenta and can only be blurredly delimited from the placenta

8

compared to the placenta and therefore does not allow a clear diagnosis in all cases. There are partly cystic and partly solid areas, which are only blurredly distinguishable from the placenta. Palpation performed simultaneously with sonography can be helpful. Under certain circumstances, floatation of the blood clots or compressibility of the hematoma is recognizable. The specificity of sonography in diagnosing premature placental abruption is 93% with a sensitivity of 28% (Glantz and Purnell 2002; Shinde et  al. 2016). Thus, premature placental abruption can be well diagnosed via B-mode ultrasonography, if visible (. Fig.  8.24). However, an inconspicuous sonographic image does not rule it out. Sonographic evidence of premature placental abruption is associated with a worse fetal and maternal outcome (Shinde et al. 2016) (. Fig. 8.25).  

 

Doppler Sonography Not every premature placental abruption leads to an emergency delivery. Sometimes the clinical picture of the patient is not clear, or one dares to wait with the delivery due to an extremely early gestational age – despite the diagnosis of a partial premature placental abruption. It is therefore natural to ask whether Doppler sonography can make a

..      Fig. 8.25  Massive retroplacental hematoma. The placenta cannot be demarcated. The sonographic image appears typically inhomogeneous

contribution to the diagnostic chain or better determine the optimal time of delivery. However, studies in this direction show that Doppler sonography is not useful in the detection, evaluation and assessment of premature placental abruption. Despite progressive detachment, normal Doppler values are regularly seen in the umbilical cord, which can probably be explained by a loss of placental resistance and the resulting barrier-­ free flow into the retroplacental space. However, a decreasing resistance in the middle cerebral artery in fetuses after the 32nd week of gestation may be the first indication of fetal asphyxia, and may support the tentative diagnosis of premature placental abruption with the inclusion of all other signs (Morales-Rosello et  al. 2017). Therefore, it remains to be stated that Doppler sonography has no value over pathological CTG in the obstetric management of premature placental abruption. However, in ambiguous situations where prolongation of pregnancy is being considered, a middle cerebral artery resistance decrease can be a valuable tool to detect the progressive fetal endangerment situation. However, the physiological decrease in resistance of the middle cerebral artery with increasing gestational age must be taken into account (Gadelha-­Costa et al. 2007).

167 Placenta-Related Hemorrhage: Pathophysiology, Diagnostics, Management

Cardiotocography (CTG) CTG monitoring is able to indicate an intrapartum change in fetal condition very sensitively. This advantage is contrasted by the low specificity of the method, which is mainly reflected in the large number of false-­positive findings. In advanced premature placental abruption, the CTG often shows abnormal heart rate patterns (tachycardia, bradycardia, restricted macrooscillation, decelerations, etc.), which indicate a fetal risk situation but are not pathognomonic. If appropriate, a hyperfrequent labor pattern may be evident. The CTG may remain classified as unremarkable for a long time even in the presence of extensive placental abruption, so that an unremarkable CTG does not rule out premature placental abruption. If the CTG already shows bradycardia when the diagnosis of premature placental abruption is confirmed (e.g. by ultrasound), this is associated in particular with severe fetal acidosis and a poor fetal outcome  – irrespective of the gestational age and the sonographic findings (Matsuda et al. 2013; Takano et al. 2013).

 agnetic Resonance Imaging (MRI) M and Computed Tomography (CT) In the clinical diagnosis of acute premature placental abruption, magnetic resonance imaging and computed tomography are not of major importance. Secondarily, however, they can be performed in the context of maternal trauma to exclude further organ damage and can be useful in clinical management. MRI is able to visualize hemorrhage as well as ischemia and inflammatory reactions in and around the placenta (Linduska et al. 2009). Visualization of retroplacental hematoma is more successful than in B-mode ultrasonography (Masselli et al. 2011). In addition to MRI, premature placental abruption in trauma patients can also be diagnosed very well on CT.  After

8

trauma, it is not uncommon for a CT to be performed to evaluate for conceivable organ damage to the mother. This can help to better assess the risk for mother and child with regard to premature placental abruption after such an event and to align further clinical management with these findings (Kopelman et  al. 2013). A grading system (Traumatic Abruptio Placenta Scale, TAPS) was published in 2014 (Saphier and Kopelman 2014). 8.4.6

Laboratory Diagnostics

 erum Markers in the First S and Second Trimester The serum markers plasma protein A (decreased), alpha-fetoprotein (increased) and inhibin A (decreased) have been discussed for the evaluation of the risk of premature placental abruption in the first and second trimester (Ananth et  al. 2017a). However, a valid screening algorithm for clinical practice does not exist (Odibo 2014). Nevertheless, the conspicuousness of serum markers in the context of maternal risk factors (age, previous diseases, etc.) is confirmed in various studies and thus forms the basis for using them in the future for risk evaluation not only in preeclampsia (Odibo 2014; Blumenfeld et  al. 2014). However, further studies are still required for this purpose.

Kleihauer-Betke Test Detection of fetal erythrocytes in maternal blood cannot be used to diagnose premature placental abruption, as a positive Kleihauer-­ Betke test does not prove premature abruption and a negative test does not rule it out (Dhanraj and Lambers 2004). A recent paper reported a sensitivity of 4.4% for the Kleihauer-Betke test in cases of premature placental abruption (Atkinson et al. 2015a).

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Blood Count/Coagulation Diagnostics

8

Maternal blood loss in premature placental abruption is reflected in the laboratory as a drop in the Hb value in the complete blood count. It is not uncommon for the blood loss to be accompanied by a coagulation disorder, which is best visible in the laboratory on the basis of fibrin cleavage products (fibrinogen level, thrombin time) and thrombocytopenia (Examination and imaging of the placenta is an important component of obstetric monitoring. The placenta should be sonographically examined and documented in all pregnant women (Merz et al. 2012).

Criteria for Assessment

Sonographic

Placental

The differentiated examination of the placenta includes the assessment of 55 Shape 55 Size and thickness 55 Echo-structure and maturity 55 Echogenicity 55 Localization 55 Tumors 55 Implantation disorders 55 Umbilical cord

The standard examination method for imaging the placenta is still sonographic assessment using 2D/3D procedures and Doppler sonography. At higher weeks of gestation and under limited examination conditions (obesity, oligo-/anhydramnios unfavorable fetal position or posterior placenta), sonographic assessment of the placenta may be limited and magnetic resonance imaging (MRI) may be helpful, especially for specific questions (Dekan and Linduska 2011). Advantages of MRI are the large field of view (FoV), soft tissue differentiation, and multiplanarity. In addition, some pathologies such as placental implantation disorders and, in some cases, infarctions and hemorrhages can be better visualized using MRI (Dekan and Linduska 2011; Jha et al. 2016). >>The normal placenta appears ultrasonographically as a homogeneous intermediate echogenic structure adjacent to the internal myometrium, from which it is separated by a hypoechogenic border zone (. Fig. 9.1).  

Depending on the MRI sequence, the placental MRI signals vary. In the T2-weighted multiplanar single-shot echoplanar turbo/ fast spin echo sequences acquired in three spatial planes (axial, sagittal, coronal), which are used as a standard technique for fetal

189 Placental Imaging

MRI examinations (e. g. HASTE, “half Fourier acquisition single shot turbo spin echo”; SSFSE, “single shot fast spin echo”; UFSE, “ultra fast spin echo” etc.; named according to manufacturer), the placenta appears as homogeneous tissue, isointense to the myometrium in early pregnancy. A thin hypointense line may represent the placenta-

..      Fig. 9.1  Normal findings in ultrasound examination

myometrium boundary. As pregnancy progresses, the placental signal becomes hypointense compared  to the myometrium. On T1-weighted sequence (“T1w spoiled gradient echo”, with frequency-­selective fat saturation or opposed phase imaging), which should be acquired in at least one spatial plane, the placenta presents hypointense to somewhat hyperintense depending on physiological changes and vascular supply (Jha et  al. 2016). The calcifications seen as the placenta ages are poorly differentiated on MRI; if large enough, they can be delineated from the surrounding area without signal. Susceptibility-weighted sequences (SWI) or diffusion-weighted sequences (DWI) with quantitative assessment of diffusivity (ADC determination, “apparent diffusion coefficient”) can be helpful in the dedicated assessment of the placenta (Bonel et al. 2010). The use of MRI contrast agents containing gadolinium is not indicated (. Fig. 9.2).  

a

b

c

d

e

f

..      Fig. 9.2  a–f Normal findings in MRI. a Anterior wall placenta in 30th week of gestation, delineate delicate hypointense (dark) borderline between myometrium and placenta; polyhydramnios (T2w Haste). b–d: Fundal placenta in  the 15th week of gestation,

9

anhydramnios and pulmonary hypoplasia in polycystic kidneys, hydrocephalus, cervical cele (b: T2w Haste, c: T1fl2d, d: T2w Haste). (e and f): 19th week of gestation with syndromal disease. Spina bifida (e: T2w, f: T1fl2d)

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This chapter is dedicated to the normal and the abnormal placenta in the second and third trimester with special attention to the sonographic appearance, the valuable use of MRI and the potential management. 9.1.2

Localization

Determination of placental localization is the most commonly performed examination of the placenta and is important for risk assessment.

9

>> While the actual placental location (anterior, posterior, fundal, lateral) is only of decisive importance in cases of low-lying, the distance of the placenta from the internal cervical os is important for obstetric management and should be assessed by vaginal sonography from the second trimester onwards (Farine et al. 1990).

Lateral placental localization may possibly lead to differences in resistance indices of the uterine arteries, with the ipsilateral uterine artery having lower indices compared to the contralateral side (Ito et al. 1990).

Low-Lying Placenta The placenta is normally located on the anterior or posterior wall of the uterus and extends to the lateral walls. The caudal edge should be at least 2 cm away from the internal cervical os; a distance of ≤2 cm, without overlapping the cervix, is called a low-lying placenta (. Fig. 9.3) (Reddy et al. 2014). Due to the stretching and growth of the lower uterine segment in the course of pregnancy, a relative “positional change” of the placenta in cranial direction can occur up to about 32 weeks gestation, this is called “cephalad placental migration.” These changes can have important consequences for clinical management, especially in the case of a lowlying placenta (Hung et al. 1999).  

..      Fig. 9.3  Low-lying placenta

The frequency of a low-lying placenta or placenta praevia depends on the gestational age: Between the tenth and 20th week of gestation, up to 6% of pregnant women present with placenta praevia (Oyelese and Smulian 2006); in the 18th–22nd week of gestation, 2–4% of all placentas reach or overlap the internal cervical os (Oppenheimer et al. 2007), and a low-lying placenta is present in 8.4% (Blouin and Rioux 2012). In 95% of placentas classified as low-lying or overlapping in the second trimester, the diagnosis was revised in the third trimester. This means that normal placental localization in the first trimester (here the anterior and posterior walls of the lower uterine segment are still adjacent) should not be misinterpreted as placenta praevia. >> If placenta praevia is “detected” in the first trimester, the diagnosis should be communicated cautiously, and follow-up examinations should be performed in the second and third trimesters (Fuchs et  al. 2008). At term delivery, approximately 0.5% of placentas reach or overlap the internal cervical os (Rosenberg et al. 2011).

However, the degree of overlap correlates with the risk of a persistent finding: if the placenta overlaps the internal cervical os 2.5 cm, the finding persists in 40% (Becker et al. 2001). A measurement of the distance between the placenta and the inner cervical os in the second trimester should be repeated again around the 34th week of gestation. Only then should a decision be made regarding the mode of delivery because of the placental location. >>A distance of the placenta from the internal cervical os ≥2 cm is not associated with an increased risk of bleeding and is considered “safe” for vaginal delivery (Oppenheimer et al. 1991, 2001).

With a low-lying placenta, the C-section rate is 31% and the likehood of obstetric bleeding is 3%. If a low-lying placenta is associated with a marginal sinus, intrapartal hemorrhage is more common. A marginal sinus is described as an area filled with maternal blood and should not be misinterpreted as vasa praevia (fetal vessels in close proximity to the internal cervical os). If the placenta is low lying, vasa praevia can be a subsequent complication with potentially serious ­consequences.

Placenta Praevia In placenta praevia, the placenta completely covers the internal cervical os (Reddy et al. 2014). According to Reddy et  al. (2014), the classification depends on the relationship to the internal cervical os (see following overview and . Fig. 9.4).  

Classification of Placental Localization (According to Reddy et al. 2014) 55 Low-lying placenta: distance to the internal cervical os >Diagnosis and thorough evaluation of a Low-lying placenta or placenta praevia by sonography and MRI, if necessary, is crucial for clinical management (Schlembach et al. 2016) because of the increased risk of the  placental accreta spectrum, umbilical cord attachment pathology and vasa praevia, and respective pregnancy complications (Hung et al. 1999; Gemer and Segal 1994).

9

In the case of a low-lying placenta or placenta praevia, the documentation/notification of the placental localization (anterior, posterior) is important for the surgical procedure, since an anterior wall placenta can lead to complications due to the transplacental delivery of the child (umbilical cord hemorrhage, difficult child development) (Fuchs et al. 2008). For the correct diagnosis of placenta praevia, both the lower placental margin and the internal cervical os must be accurately identified, if necessary also by means of transvaginal ultrasound. This is all the more important as, in the case of placenta praevia, primary C-section must be performed between the 36th–37th week of gestation before the onset of labor. However, ultrasound is still often inaccurate and incorrect in the diagnosis of ­placenta praevia. Placenta praevia can be mistaken for a low-lying placenta, which overlaps the internal cervical os but which has not implanted into the opposite segment. Color Doppler can be used to distinguish between truly implanted and non-implanted placental tissue. Another source of error is the distinction between the lower uterine segment and the cervix. The uterine isthmus is usually closed during early pregnancy and only opens at higher gestational age. Due to the improved resolution of newer ultrasound machines, recent studies have been able to address the distinction between these two structures and assess the length of the cervix and isthmus

(Greco et  al. 2011; Souka et  al. 2011; Hasegawa et al. 2017). A reliable diagnosis of placenta praevia is made using transvaginal ultrasound between the 20th and 24th week of gestation, after opening of the lower uterine segment and by accurately distinguishing between the cervix and isthmus (. Figs. 9.5, 9.6, and 9.7) (Hasegawa et al. 2017). Other causes of misinterpretation are a full bladder pressing on the lower uterine segment and segmental uterine contractions.  

..      Fig. 9.5  Placenta praevia

..      Fig. 9.6  Placenta praevia. T2-weighted HASTE sagittal—the placenta covers the cervix incompletely in the anterior wall region (32nd week)

193 Placental Imaging

a

9

b

..      Fig. 9.7  a, b Placenta praevia. T2-weighted sequence sagittal a with displacement of the placenta in front of the cervix. The basal decidual membrane to

the myometrium appears intact. T1-weighted sequence b with hyperintense mucus plug and irregular placental hemorrhage

>> In all pregnant women, the placental location should be examined sonographically and, in case of a low-lying placenta, the presence of vasa praevia should be examined and documented, if necessary as part of a further ultrasound examination (Merz et al. 2012; Schlembach et al. 2016).

Untypical Placental Location

This is also possible by abdominal sonography in case of a low-lying placenta with a sufficiently filled urinary bladder. Tip

If placenta praevia is suspected, vaginal ultrasonography should be performed with an empty bladder (Heer et al. 2006). In contrast to digital examination, vaginal sonography is not contraindicated and does not lead to an increased risk of bleeding (Tikkanen et  al. 2006). For a possible supplementary MRI, slice arrangements oriented sagittally to the vagina are recommended, as they best reveal the positional relationship of the placenta to the cervix.

Usually no clinical distinction is made between the exact location of anterior, lateral or posterior placenta. However, certain localizations are thought to be associated with certain fetal positions and attitudes, such as a breech presentation or an occiput posterior position with an anterior wall placenta (Fianu and Vaclavinkova 1978; Gardberg and Tuppurainen 1994). Recently, studies showed that a placenta that is neither anterior nor posterior in the second trimester is associated with an increased incidence of adverse obstetric outcomes (Fung et  al. 2011; Seckin et al. 2015). 9.1.3

 lacenta Accreta Spectrum P (PAS)

As a consequence of a partial or total absence of the maternal decidua, which leads to direct contact between the chorion frondosum and the myometrium, there may be disturbances in placental implantation with abnormal adherence of the placental villi to the maternal myometrium.

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Depending on the extent and depth of invasion of the placenta, different implantation disorders are distinguished (see following overview) (Belfort 2010). Classification of Placenta  Accreta Spectrum (PAS) (Belfort 2010) 55 Placenta accreta: invasion up to the myometrial inner wall (81.6%) 55 Placenta increta: invasion into the myometrium (6.6%) 55 Placenta percreta: invasion to the uterine serosa or beyond the uterine borders (11.8%)

9

PAS occurs in 0.9% of all pregnancies and in 9.3% of pregnant women with placenta praevia. The risk of PAS increases with the number of cesarean deliveries (. Table 9.1).  

>>Eighty to ninety percent of all PAS present with placenta praevia (Garmi and Salin 2012). Additional predisposing factors are previous uterine surgeries (curettage, enucleation of fibroids), submucous fibroids, endometritis and placental abruption in the medical history (Garmi and Salin 2012).

Imaging All pregnant women at increased risk for PAS should receive prenatal assessment for placental implantation (Merz et  al. 2012; Schlembach et al. 2016). Diagnostic imaging includes sonographic assessment and MRI examination when appropriate. Prenatal detection of PAS is possible in approximately 50–80% (Soto and HernándezAndrade 2015), and clinical suspicion should be followed by MRI examination— suspicion of PAS is the main indication for MRI examination of the placenta (Dekan and Linduska 2011; Soto and HernándezAndrade 2015). The lower uterine segment close to the cervix, a deep placental infiltration under the bladder and a possible cervical invasion in the case of placenta praevia can be examined more precisely by vaginal sonography than by abdominal sonography. Sonographic assessment is facilitated by a filled bladder in both approaches, as the bladder filling provides an ideal ultrasound window and the bladder wall is unfolded (Fuchs et al. 2008). Three-dimensional imaging of the findings can provide further hints, especially for ruling out the possibility of the placenta penetrating into the surrounding structures.

..      Table 9.1  Frequency of an abnormal placental implantation in relation to the number of previous C-sections and placenta praevia (Silver et al. 2006) Cesarean section

PAS (%)

OR (95% CI)

Without placenta praevia (%)

With placenta praevia (%)

1 (primary)

0.2

–

0.03

3.3

2 (condition after C-section)

0.3

1.3 (0.7–2.3)

0.2

11

3

0.6

2.4 (1.3–4.3)

0.1

40

4

2.1

9.0 (4.8–16.7)

0.8

61

5

2.3

9.8 (3.8–25.5)

0.8

67

≥6

6.7

29.8 (11.3–78.7)

4.7

67

195 Placental Imaging

In the case of posterior wall placenta, the assessment of placentation is difficult due to the poorer ultrasound window without the urinary bladder as an anterior route or due to obstruction of the view by the fetus. Especially in these cases, the MRI examination can provide valuable information as an additional diagnostic procedure. Sonographic Signs of PAS PAS  should be considered with the following findings (. Figs.  9.8 and 9.9) (Fuchs et al. 2008; Belfort 2010): 55 Multiple lacunae in the placenta (“Swiss cheese”) 55 Loss of the hypoechogenic retroplacental zone 55 Blood vessels or placental tissue break through the uteroplacental boundary  

a

9

55 Blurred demarcation between placenta and myometrium with retroplacental myometrium thickness < 1 mm 55 Hypervascularization

Placental lacunae with turbulent flow  (vascular bridges extending from the placenta into the myometrium) are the most characteristic sign of PAS with a sensitivity of 78– 93%. The more lacunae to be visualized, the more likely an invasion disorder is present (Soto and Hernández-Andrade 2015). In contrast, the loss of the hypoechogenic retroplacental zone has a sensitivity of only 7% (Soto and Hernández-Andrade 2015). Even though the diagnostic value of MRI has not yet been adequately clarified (Fuchs et  al. 2008; Belfort 2010), an MRI examination should usually be performed in the third trimester in the case of sonographic b

..      Fig. 9.8  a, b Placenta accreta spectrum (PAS). a Multiple lacunae (“Swiss cheese”) and loss of the hypoechogenic zone; b Hypervascularisation

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Urinary bladder

Urinary bladder

Placenta Placental vessel Placental vessels

..      Fig. 9.9  Placenta accreta spectrum (PAS). Blood vessels break through the uteroplacental boundary to the urinary bladder

9

evidence of PAS in order to plan the delivery. The urinary bladder of the pregnant woman should be moderately full in order to better assess the bladder wall and to differentiate varices of the bladder wall from invasion or to avoid overestimation of invasion if the bladder is too full. It is essential that the MRI images are interpreted with knowledge of the sonographic examination/ results and discussed jointly by radiologists and obstetricians. The large field of view of MRI makes it possible to visualize and assess the entire placenta. A detailed description of the structures involved (bladder, bowel, pelvic wall) is important for surgical planning (e.g. insertion of a ureteral stent). MRI Criteria of PAS The diagnostic criteria on MRI are similar to the sonographic features (Soto and Hernández-Andrade 2015; Lax et  al. 2007; Bardo and Oto 2008): 55 Uterine bulging: “protrusions” of the uterus 55 Heterogeneous inhomogeneous signal infiltrating from the placenta into the myometrium

55 Irregularly configured, hypointense lacunae in T2 weighting (blood flow/ vascularization)—“Swiss cheese” 55 Hypointense intraplacental bands in T2 weighting 55 Focal invasion of the myometrium

According to Lax et al. (2007), some criteria important for sonography, such as thinned retroplacental myometrium, visualization of an exophytic mass and an  irregular hyperechogenic border between uterus and bladder, are not useful for assessment by MRI. A thinning of the myometrium or a tenting in the area of the urinary bladder are unspecific signs, which should, however, draw attention to look for further changes. In addition, a possible invasion into the parametria has to be evaluated. A placenta percreta can be described relatively reliably by MRI.  Less pronounced forms of invasion of the placenta (accreta, increta) are more difficult to detect and differentiate (. Figs. 9.10 and 9.11); however, this does not tend to influence the further procedure and the surgical technique. It must also be  

197 Placental Imaging

borne in mind that different manifestations of invasion can occur in a pregnancy. A mere bulging of the uterus into the urinary bladder does not prove the presence of a placenta percreta, since the myometrium is thinned out and altered by scarring, especially in the case of previous C-section. Therefore, there is a risk of false-positive overestimation of invasion. In these cases, it

9

is essential to demonstrate T2-weighted hypointense lacunar vascular connections between the uterus and the urinary bladder in order to prove the depth of invasion before surgery (. Fig.  9.12). The intraplacental lacunae impose hypointense in the T2-weighted image; bizarre and irregular configurations allow differentiation from the typical placental septa formed during formation of the cotyledons. According to a meta-analysis, sonography and MRI in principle have comparable predictive value with regard to PAS (D’Antonio et al. 2014).  

9.1.4

Echogenicity

The normal placenta appears relatively homogeneous. There are different types of abnormal echogenicity. Hypoechogenic lesions are the most common placental abnormalities and are usually diagnosed after the 25th week of gestation.

..      Fig. 9.10  Placenta increta. T2-weighted sagittal MRI with clearly hypointense caverns. The boundary to the myometrium is only clearly recognizable at the anterior wall

a

..      Fig. 9.11  a, b Placenta increta. T2w sagittal a with placenta praevia infiltrating the myometrium. Evidence of thick vacuolar lacunae in anterior wall

>>When examining the placenta, the complete placenta must be assessed in order to exclude placental lesions. If lesions are visualized, a Doppler sonographic examination should be followed.

b

region. Boundary to urinary bladder irregular, a breakthrough cannot be verified even in magnification b

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a

b

c

..      Fig. 9.12  a–c Placenta percreta. Evidence of T2w hypointense vascular structures in the border region towards the urinary bladder. These are best delineated in the T2-weighted sequence with fatty saturation, whereas in the T1w the vessels cannot be identified.

9

Within the placenta in all sequences inhomogeneous signal and tubular vascular structures, the T1w hyperintense areas are most suggestive of fibrous/scar tissue in the placenta. a T2w HASTE; b TIRM; c T1w GRE

Placental Lacunae Placental lacunae represent collections of maternal blood in the intervillous space (Kanne et al. 2005) and are the most common form of hypoechogenic areas (usually >2 cm) without villous structures. They result from reduced intervillous blood flow, thrombosis, or fibrin deposition (Dekan and Linduska 2011) and are most commonly found in the late second and third trimester. The characteristic swirling venous flow can be visualized in real time. Placental lacunae (. Fig.  9.13) should not be mistaken by the irregular, vascular lacunae typical for adherent placenta. Placental lacunae occur during pregnancy in 20–67% of all pregnancies (Bowman and Kennedy 2014) and have no clinical significance (Soto and Hernández-­Andrade 2015; Thompson et al. 2002). There are casuistics of MRI examinations  reporting T1-hypointense and T2-­ isointense signal behavior—comparable to fresh blood (Morikawa et al. 2005).  

..      Fig. 9.13  Thick jelly-like placenta with placental lacunae (asterisks)

Septal Decidual Cysts Septal cysts of the decidua (. Fig.  9.14) arise from focal degeneration within the  

..      Fig. 9.14  Septal decidual cysts

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maternal decidua. They are found in 20% of placentas at term. Septal cysts represent another form of hypoechogenic lesion, are usually 3 cm, especially near the base of the umbilical cord, regular biometry should be performed and fetal cardiac function should be monitored (Raga et al. 1996; Brown et al. 2002).

 chogenic Cystic Lesions (Placental E Bed Infarction) These lesions, 1–2  cm in diameter, have a central fluid-filled space and usually present clearly (. Fig.  9.15). Echogenic cystic lesions can be distinguished from other lesions by their hyperechogenic margin, which represents perivillous fibrin deposi 

..      Fig. 9.15  Echogenic cystic lesions

9

tion, and their irregularly notched border. In addition, unlike placental lacunae, there is no flow on color Doppler. The etiology of echogenic cystic lesions is unclear. Approximately 10–15% of all pregnant women with pregnancy complications, or 0.09% of all pregnant women form pronounced perivillous fibrin deposits, so-called Gitter infarcts. According to recent research, these are intervillous thrombi, which consist of coagulated maternal blood in the intervillous space surrounded by compressed or infarcted villi (Soto and Hernández-Andrade 2015; Harris et  al. 1990, 1996). They are associated with adverse pregnancy outcome (habitual abortions, IUFD, FGR, preeclampsia, preterm delivery) (Proctor et  al. 2010). The triad of echogenic cystic lesions, abnormal uterine artery Doppler and abnormal ­placental shape is usually indicative of severe placental impairment and can predict perinatal death with a probability of 52% (Viero et al. 2004).

Placental Infarction Placental infarctions are detectable in approximately 20% of all uncomplicated pregnancies and in 40% (of mild) and 70% (of  severe) preeclampsia (Moldenhauer et al. 2003; Krielessi et al. 2012). In 39% of all pregnant women with preeclampsia, the placenta is more than 5% infarcted (Vinnars et al. 2011). Placental infarctions present as echogenic or even anechogenic avascular areas and result from lack of dilatation or thrombotic occlusion of the spiral arteries, increased perivillous fibrin deposition, and disruption of fetal circulation due to fetal thrombotic vasculopathy (Soto and Hernández-­Andrade 2015). On MRI, ischemic infarctions of the placenta can be delineated as hyperintense in T2 weighting and diffusion weighting (DWI) trace image; hemorrhagic and thrombotic infarctions are hyperintense in T1 weighting (Linduska et al. 2009).

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Placental Hematoma Hematomas present sonographically as hypo- to anechogenic (acute or chronic) preplacental (to the fetus), retroplacental (to the mother; . Fig.  9.16), retrochorial or retroamniotic zones distributed marginally along the placental border or intraplacentally. Subacute isoechogenic hemorrhage can be verified by displacement effects, for example, on perfusion assessment. On MRI examination, hematomas may have variable signal intensities, depending on the age and composition of the hemorrhage (. Fig.  9.17). Subacute hemorrhages should be better differentiated by MRI tomography than by sonography; however, any therapeutic decision will be determined by clinic features and not by MRI (Masselli

et al. 2011). In T1 imaging, hematomas can be delineated hyperintensely, whereas in the more T2-weighting blood-­sensitive gradient echo (GRE) sequence, they are hyopinten-

 

 

9

..      Fig. 9.16  Retroplacental hematoma  (11.2  cm × 9.5 cm)

a

b

c

d

..      Fig. 9.17  a–d Placental hematomas/hemorrhages on MRI. a, b T1w images (T1 fl2d): the hyperintense changes within the inhomogeneous placenta correspond to hemorrhages (a: coronary, b: sagittal). c, d

T2w and T1w axial images: T2w-hypointense and T1w-­hyperintense circumscribed changes, at 36th week of gestation with circumscribed hemorrhages within the placenta (c: T2 Haste tra; d: T1 fl2d tra)

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9

.       Table 9.2  Ischemic and hemorrhagic pathology on MRI (Dekan and Linduska 2011) T1w

T2w

GRE

DWI

Localization

Morphology

Ischemia

−

+

−

+

Intraplacental

Diffuse/delineated

Hemorrhage

+

−

+

+

Intraplacental

Diffuse/delineated

Subchorial hemorrhage

+

−

+

+

Subchorial

Delineated

Intervillous hemorrhage

+

−

+

+

Intraplacental

Round

Intervillous thrombi

−

+

±

±

Intraplacental

Delineated

Retroplacental hematoma

+

−

+

+

Retroplacental

Delineated

+ hyperintens, − hypointens, GRE gradient echo, DWI diffusion weighting

sive (. Table  9.2) (Dekan and Linduska 2011). Randomly verified blood collections without clinical symptoms are mostly venous in origin and should only be  controlled by follow-up examinations. Hematomas are associated with adverse pregnancy outcomes (habitual abortions, IUFD, placental abruption, FGR, preterm birth), but lack of uniform standardized definition criteria makes it difficult to make a valid statement about the incidence of pregnancy complications (Soto and Hernández-­Andrade 2015).  

Breus’ Mole

..      Fig. 9.18  Jelly-like placenta

It is a rare, distinct (>50% of placental area) subchorial hemorrhage of unclear etiology, first described by Breus in 1892 (Jha et  al. 2016). Due to massive hemorrhage, there is a high risk of adverse pregnancy outcome (FGR and in up to 50% IUFD) (Alanjari et al. 2013). The sonographic picture is identical to that of hematomas: Initially, hyperechogenic areas without blood flow are seen, which take on a heterogeneous appearance with increasing clot formation.

Jelly-like Placenta The jelly-like placenta is characterized by inconsistent echogenicity and anechogenic spaces (. Figs. 9.18 and 9.19). It moves like  

..      Fig. 9.19  Thickened jelly-like placenta, anhydramnion

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jelly in response to abdominal pressure (Jauniaux et al. 1990). Placental thickness is usually >95th percentile (Jauniaux et  al. 1994). Most jelly-like placentas are located laterally, close to the fundus (Raio et  al. 2004). This location is often associated with poor perfusion, preeclampsia and FGR (Kofinas et al. 1989). Jauniaux et al. were the first to describe the triad of abnormal maternal blood counts, pathological uterine Doppler and small j­elly-­like placenta with anechogenic lakes in FGR and hypertension (Jauniaux et al. 1994).

Mesenchymal Dysplasia of the Placenta

9

Placental mesenchymal dysplasia was described in 1991 as a placental vascular abnormality with diffuse hyperplasia of the villi (Moscoso et al. 1991). With a thick placenta displaying hypoechogenic areas, the ultrasound findings characteristic of this entity are similar to those of a hydatidiform mole (. Fig. 9.20). However, in contrast to partial moles, most fetuses with placental mesenchymal dysplasia have an unremarkable karyotype, and ßhCG levels remain normal throughout pregnancy. In order to prevent non-indicated abruptions, placental mesenchymal dysplasia must be clearly distinguished from hydatidiform moles (Parveen et al. 2007). If a sus 

..      Fig. 9.20  Mesenchymal dysplasia of the placenta

pected diagnosis of placental mesenchymal dysplasia is made after ultrasound diagnosis and genetic testing, the affected patients must be treated as high-risk pregnant women, since placental mesenchymal dysplasia is associated with fetal growth restriction, intrauterine fetal death and other chromosomal abnormalities. 9.1.5

Maturation of the Placenta

Sonographically, the degree of maturity of the placenta can be divided into four grades according to Grannum et al. (1979) (see following overview; . Fig. 9.21).  

Sonographic Classification of Placental Maturation (According to Grannum et al. 1979) 55 Grade 0: homogeneous parenchyma, smooth bordered chorionic plate (approx. 12–30 weeks of gestation) 55 Grade I: slightly wavy chorionic plate, single parenchymal  echo enhancements (approx. 30–32 weeks of gestation) 55 Grade II: elongated echo enhancements from the chorionic plate towards the basal plate, echo-rich structures in the parenchyma (32–35 weeks of gestation) 55 Grade III: chorionic plate interrupted (compartmentalization), cotyledons recognizable, hyperechogenic structures in the parenchyma, garland-­ shaped pattern (>36th weeks of gestation)

The degree of maturity of the placenta is based primarily on the presence and distribution of calcifications from calcium deposits. In the late 1970s, Grannum et  al. developed a method for determining placental maturity to predict fetal development (Grannum et  al. 1979). According to this method, Grannum 0 corresponds to a less

9

203 Placental Imaging

Grade 0

Grade I

Grade II

Grade III

..      Fig. 9.21  Sonographic representation of placental maturity grades 0–III

mature placenta, whereas Grannum III corresponds to a very mature placenta. The assessment of placental echogenicity (decreased or increased) can be used in the identification of risk constellations (Jauniaux et al. 1990; Raio et al. 2004). >>Analogous to the sonographic classification according to Grannum, MRI can also determine the degree of maturity of the placenta. T2-weighted sequences show the placenta in early pregnancy to be moderately hyperintense and thus well demarcated from the myometrium. If the placenta is relatively homogeneous in T1 and T2 weighting in the first and early second trimester, it becomes inhomogeneous with irregular surface and lobulated internal structures (cotyle-

dons) in the further course (Dekan and Linduska 2011; Blaicher et al. 2006).

However, the assessment of placental maturity has lost importance in recent years due to the weak correlation with a poor perinatal outcome and especially due to the high subjectivity of the method (Sau et al. 2004; Moran et al. 2011), especially since Doppler sonography is a much better method for assessing the fetoplacental unit. 9.1.6

Size and Shape

The placenta is usually round to oval with a wide variability (Yampolsky et  al. 2008). Irregular shapes are mainly determined by localization, atrophy and implantation.

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 ccessory Placenta (Placenta A succenturiata) In about 8%, an accessory lobe (placenta succenturiata) is present, which is connected to the main placenta by vascular bridges and caused by partition. The most common variant is placenta bilobata (Meizner et  al. 1998; Soto and Hernández-Andrade 2015). A placenta succenturiata can be the cause of fetal hemorrhage (due to rupture of vasa aberrantia—especially during rupture of the fetal membranes) or of placental retention (Suzuki and Igarashi 2008). Therefore, a careful search for aberrant vessels (especially vasa praevia) should be carried out, especially in the case of accessory placentas in the lower uterine segment.

9

>>Accessory placentas should always be excluded sonographically, as they may remain in utero and thus lead to postpartum complications (bleeding).

Placenta circumvallata Placenta circumvallata is a morphologically abnormal placenta in which the transition from membranous (chorion laeve) to villous chorion (chorion frondosum) is not at the margin of the placenta but at some distance from it (. Fig. 9.22) (Scott 1960). The chorionic plate on the fetal  

..      Fig. 9.22  Placenta circumvallata

side, from which the villi originate, is smaller than the basal plate on the maternal side of the placenta. In this constellation, placental tissue is located outside the boundary of the chorionic plate, giving rise to the name “placenta extrachorialis.” A double layer of amnion and chorion with necrotic villi and fibrin forms an annular placental rim. Difficult to detect prenatally, one may see sonographically irregular, raised placental margins as well as a peripheral ring of chorionic tissue, which appears as an echodense ridge (Soto and Hernández-Andrade 2015; McCarthy et al. 1995; Harris et al. 1997; Suzuki 2008; Elsayes et  al. 2009). In three-­dimensional ultrasound, this feature appears like a tire (the “tire sign”) (Arlicot et  al. 2012). The incidence of placenta circumvallata is 1–2%, and it is associated with an increased rate of perinatal complications, such as prematurity, oligohydramnios, premature rupture of membranes, pathological CTG, placental abruption, and intrauterine fetal death (IUFD).

Placenta membranacea/Placenta diffusa Placenta membranacea/placenta diffusa represents a rare placental anomaly (incidence approximately 1/20,000 births, 1/185 for placenta praevia) (Soto and HernándezAndrade 2015), possibly associated with placenta accreta (Pereira et  al. 2013). The amnion is completely or partially covered with a thin membrane of villous tissue (Dekan and Linduska 2011; Soto and Hernández-Andrade 2015). The risks reported here are pre- and postpartum hemorrhage, late abortions, and fetal growth restriction (FGR) with an increased risk of IUFD (Soto and Hernández-­ Andrade 2015). Placenta membranacea should be considered if imaging shows a thin layer of placental tissue over most of the uterine cavity.

205 Placental Imaging

9.1.7

Placental Biometry and Volumetry

Biometry and volumetry of the placenta have been suggested as potentially useful methods for estimating perinatal risks (Elsayes et al. 2009). For area and volume calculations, a circular placental shape is generally assumed. However, in conventional ultrasound examination, two-dimensional measurements (thickness, diameter of the placenta) and area calculations derived from them can be achieved with significant variability of the measured values. Placental volume can also be determined using 3D sonography and MRI.  Sonographic volume calculation is performed either by means of multiplanar measurements or special algorithms such as VOCAL (Virtual Organ Computer-Aided Analysis) (Kalache et al. 2003). The diameter of the placenta is approximately 18–20 cm, and the normal placental thickness is approximately 2–4  cm (Kaplan 2008; Lee et al. 2012) measured centrally or at the base of the umbilical cord if it is centrally located. In daily clinical routine, the placental thickness is only estimated subjectively and, if it appears inconspicuous, not measured. Tip

It is recommended to measure placental thickness  at the site of  placental cord insertion.

Placental location and, if possible, gestational age must be considered when evaluating placental thickness. If the umbilical cord insertion is approximately central (90% of placentas are circular and have a central umbilical cord insertion), this will provide a correct measurement in most cases. If the umbilical cord insertion is marginal, this method will often measure a placenta that is too thin (Lee et al. 2012).

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In normal pregnancy, the placenta shows a continuous growth in thickness until about the 37th week of gestation, whereat the thickness of the placenta (in mm) corresponds to the gestational age in weeks (Schlensker 1971). Anterior wall placentas are about 0.7 cm thinner than posterior wall and fundal placentas. Anterior wall placentas in the second trimester of >3.3 cm thickness  and posterior wall placentas >4.0  cm thickness  are considered “abnormal” (Soto and Hernández-­Andrade 2015; Lee et  al. 2012; Hoddick et al. 1985), and a thickness > 5 cm is considered pathological (Elchalal et al. 2000). Placentas that are too thick, as well as placentas that are too thin, indicate an increased perinatal risk as non-specific findings (Jauniaux et al. 1994). Postpartum examination of placentas in case of extreme preterm birth with FGR revealed that more than 50% have abnormalities in size, shape, and umbilical cord insertion (Pomorski et al. 2012; Walker et al. 2012). Studies of placental morphology show that placentas from women with preeclampsia tend to be oval rather than round and have reduced surface area (Burton et al. 2010). Marginal and velamentous cord insertions are associated with smaller placentas and smaller newborns (Vinnars et al. 2011).

Thick Placenta Several studies described an association between increased placental volume and adverse pregnancy outcomes such as placental abruption, FGR, hypertension, neonatal acidosis and fetal death (Jauniaux et  al. 1994; Raio et al. 2004; Elchalal et al. 2000; Eskild et al. 2009; Dombrowski et al. 1992; Proctor et al. 2009; Cooley et al. 2011; Porat et  al. 2013; Miwa et  al. 2014). However, at the time of diagnosis of a thick placenta, fetal and maternal blood flow, an early sign of abnormal conditions, was not different between the two groups (Arabin et al. 1992). This suggests that the ultrasonographically

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..      Fig. 9.23  Thick placenta

detected thick placenta (. Fig. 9.23) corresponds to the latent phase of a placental dysfunction. A possible explanation for the increase in size due to placental dysfunction could be the compensatory proliferation and edema of the placental villi (Raio et al. 2004; Fox and Elston 1978; Todros et  al. 1999a). In other words, the thick placenta would be the result of compensatory hyperplasia of certain placental areas that are not affected by inadequate uteroplacental blood flow. Another possible explanation could be the loss of anchoring villi. In a normally developed placenta, the distal tips of the placental villi are anchored to the decidua, providing a solid structure, so that pressure in the maternal vessels cannot affect the surface of the fetal chorionic plate. This is consistent with the observation that a proportion of thick placentas are associated with a marked arrest in placental development, which is characterized by hypoplasia of distal villi (. Fig.  9.24) (Macara et al. 1996). The hypoplastic villi are the first sign of a disturbance in the formation of the villi responsible for gas exchange when angiogenesis has stopped (Macara et  al. 1996). Areas with fewer villi are therefore filled with maternal blood and present on sonography as slowly moving even beyond the  

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basal plate. These placentas appear to shrink after birth as greater volumes of maternal venous blood likely flow out of the intervillous space via the disrupted uteroplacental veins (Porat et al. 2013). Nevertheless, three-dimensional placental volume measurement cannot be used to screen for FGR. In high-risk cases, the placental volume consists mainly of maternal blood and not of functional placental villous tissue. In this respect, the sole assessment of placental size and shape as well as echogenicity by 2D sonography is more informative for serious abnormalities in placental development.

Small Sized Placenta In sonographic volume assessment, the main limitation of the method is the increasing size of the placenta with increasing gestational age and the change in the shape of the pregnant woman’s abdomen. From the late second trimester onwards, it is difficult to capture the complete placenta with the volume transducer (Hata et al. 2011); hence, the scientific focus of placental volumetry has been on the value of the method in detecting or predicting pregnancy complications. Small placentas are associated with perinatal complications. A prospective study of 712 women showed that low placental volume in the second trimester precedes low birth weight. This association already exists in the first trimester, which could be shown by means of 3D volume measurement (Hafner et al. 2001a, b). In particular, the working group around Hafner investigated the association of placental volume in the first trimester and found a significant association with preeclampsia developing later (Hafner et  al. 2006). Other research groups (Odeh et  al. 2011a; Odibo et al. 2011; Rizzo et al. 2012) were unable to establish an association between placental volume and pregnancy complications, so that ultimately the determination of placental volume as a prognos-

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a

c

(1) Normal placenta

b

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(2) Loss of chorion

(3) Increase in placental thickness

Placental insufficiency

..      Fig. 9.24  a–c Scheme for the development of a thick placenta

tic marker for pregnancy complications has not been established in clinical practice.

Placental Abruption >>The diagnostic value of an ultrasound examination for premature placental abruption is limited; the diagnosis is usually made clinically (abdominal pain, contractions, and possibly bleeding).

A detachment of >50% increases the risk of IUFD (Soto and Hernández-Andrade 2015). Ultrasound or MRI may be relevant for classification of location (subchorial, retroplacental, retroamnial) in less pronounced hematomas/detachments (Nyberg et al. 1987).

The acute hemorrhage presents sonographically hyperechogenic compared to the surrounding placental tissue. With increasing age of the hematoma the area becomes increasingly hypoechogenic (Nyberg et al. 1987). 9.1.8

Summary

Ultrasound examination of the placenta is increasingly used to detect high-risk pregnancies. Several studies show a clear association between a sonographically abnormal placenta and poor perinatal outcome. Abnormal placental morphology may imply altered placental structure, which may lead to poor perinatal outcome.

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Doppler Sonography/ Functional Diagnostics

Dietmar Schlembach

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The placenta supplies the fetus with nutrients. As an endocrine organ, the placenta produces a large number of hormones that influence the mother and the fetus, but also placental development itself. The proper formation and development of the placenta and the utero- and fetoplacental vascular systems are thus essential for adequate placental function, especially the supply of oxygen and nutrients to the fetus. Disturbance of placental function (including disturbance of utero- or fetoplacental perfusion) can lead to complications resulting in increased maternal and perinatal morbidity and mortality (fetal growth restriction [FGR], intrauterine fetal death [IUFD], preeclampsia, fetal malformations) (Graf 2008; Pasca and Penn 2010). The function of the placenta can be disturbed by various mechanisms (e.g. disturbance of gene expression, infections, premature birth) with immediate maternal and fetal/neonatal consequences and possible consequences in the long-term outcome (Pasca and Penn 2010). >>The gold standard for evaluation of the utero- and fetoplacental circulation is Doppler sonography.

MRI scans can also be used to visualize vascular placental pathologies (Messerschmidt et  al. 2011), but this is not currently of any value in clinical practice. >>The basic prerequisite for a sufficient evaluation of the findings is an exact knowledge of the morphology, physiology and pathology of the placenta (7 Chaps. 3 and 4) as well as of the feeding and draining vessels.  

9.2.1

Placental Vascular System

With two circuits—uteroplacental and fetoplacental—the placenta is an extremely complex organ that reaches its maximum functional capacity before birth. Disturbances in normal development can have a drastic effect on fetal well-being. Over the course of nine months of development, the placenta forms a vascular network approximately with a length of 500 km and a surface of 12–14 m2 through vasculogenesis and angiogenesis (Burton and Fowden 2015). Control of this vascular network and placental circulation occurs in the absence of autonomic nervous regulation through the release of local factors. The muscular arterial vessels of the villi constitute the primary resistance flow area of the placenta, the tone of which is modulated by nitric oxide (NO), among other factors (Myatt 1992). Through this complex regulation, blood flow in the placenta is matched to maternal perfusion to ensure optimal placental function (Burton and Fowden 2015). The placenta is characterized by a special cell type—the trophoblast. The trophoblast differentiates into the villous and extravillous trophoblast. The villous trophoblast forms the villous tree of the placenta, while the extravillous trophoblast invades the placental bed and stimulates remodeling of the spiral arteries, and is thus crucial for adequate uteroplacental blood flow and fetal delivery of oxygen and nutrients (Burton and Fowden 2015; Sheppard and Bonnar 1981; Everett and Lees 2012; Osol and Moore 2014). Utero- and fetoplacental perfusion depend on blood pressure, vascular resistance and blood viscosity. Various factors, e.g. drop of blood pressure, changes in vascular tone, vascular/endothelial lesions or placental infarctions, intervillous thrombosis and placental hematoma can affect uteroplacental perfusion. Fetoplacental perfusion may be affected, for example, in labor (con-

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striction of vascular lumen and increase in resistance), thrombosis or disruption of the degree of maturation of the villi, and pathological changes in the placenta (Giles et  al. 1985; Voigt and Becker 1992; Hitschold et al. 1993; Krebs et al. 1996; Todros et al. 1999ab; Shilling et al. 2014; Baron et al. 2015).

Fetoplacental Hemodynamics Via the umbilical vein, 20–30% of the oxygenated and nutrient-rich blood reaches the liver and 70–80% reaches the heart via the ductus venosus (Chaoui et al. 2014) and the fetal circulation. From there, the fetal oxygen-­depleted blood flows back into the placenta via the umbilical arteries. >>The blood flow in the umbilical arteries is a measure of the size of the perfused fetoplacental vascular tree.

The larger the vascular tree, the lower the vascular resistance, i.e. the greater the blood flow. The size of the fetoplacental vascular tree (fetal intravillous blood volume) is the product of villous vascularization and placental weight or volume (Graf 2008; Giles et al. 1985; Hitschold et al. 1993; Krebs et al. 1996; Todros et al. 1999ab). With normal placental histology (vascular tree and villi) and normal placental weight, unremarkable blood flow patterns can be visualized in the umbilical arteries unless additional factors are present that impair perfusion, such as compression of the umbilical cord (Graf 2008).

Uteroplacental Hemodynamics The uterine arteries arise from the internal iliac arteries and divide at the level of the inner cervical os into an ascending and a descending artery. From the ascending arteries, on the lateral wall of the uterus, arise the arcuate arteries, which form a vascular network with vessels of the contralateral side and from which arise the radial arteries. The radial arteries pierce the myometrium and

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divide into basal arteries and spiral arteries at the junction with the endometrium or decidua. The spiral arteries pass through the decidual basal plate (Graf 2008). Before the eighth week of gestation, invasive endovascular (now: endoarterial) ­trophoblastic plugs occlude the spiral arteries, while minimal hemochorial perfusion exists (Jauniaux et al. 2000), and the embryo is supplied by “histiotrophic” nutrition— secreted by uterine glands (Burton et  al. 2002). As gestational age advances, the spiral arteries open and intervillous circulation is complete by the end of the first trimester (Burton and Fowden 2015; Burton et  al. 2010). Maternal blood reaches the intervillous space via the spiral arteries of the uterine arteries, circulates around the chorionic villi, and drains peripherally via sinusoidal veins in the decidual septa and via the marginal sinusoids in the placental margin (Graf 2008; Burton and Fowden 2015). Remodeling of the spiral arteries results in low resistance wide vessels that carry large volumes of blood from the maternal vessels into the intervillous space without high vascular resistance (Osol and Moore 2014; Burton and Fowden 2015; Redman and Sargent 2005). Failure to remodel the spiral arteries results in altered perfusion of the placenta and fetus and an increased incidence of pregnancy complications (FGR, preeclampsia) (Redman and Sargent 2005).

Disorders of Uteroand Fetoplacental Blood Flow A large number of studies have demonstrated the association of placental insufficiency and pathological utero- and fetoplacental flow patterns. These changes are closely associated with pregnancy complications and adverse neonatal outcome and often proceed in a temporary cascade (Ferrazzi et  al. 2002; Baschat et  al. 2007). The vessels important for diagnosis and evaluation are first of all the uterine arteries and the umbilical arteries, as these vessels

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represent the maternal and fetal circulation, respectively. Disturbances in placentation are reflected in increased resistance indices in the uterine and/or umbilical vessels (. Figs. 9.25 and 9.26).  

>> Impaired uteroplacental perfusion is diagnozed when the mean pulsatility index (PI) is above the 95th percentile and/or a persistent notch is present.  It significantly increases the risk of placental dysfunction in both low- and high-risk collectives (Hernandez-Andrade et al. 2002; Cnossen et al. 2008; Gómez et al. 2008).

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At term, increased resistance in the uterine arteries is associated with an increased risk of adverse perinatal outcome, independent of fetal weight (Monaghan et al. 2018). However, the presene of an impaired uteroplacental perfusion does not allow to distinguish between the different placenta-associated pregnancy complications (preeclampsia, placental abruption, FGR, IUFD), but aids in the differential diagnosis of FGR (Kehl et al. 2017). Estimation of the uteroplacental resistance in first and second trimesters is currently being promoted as a potential screening method for the detection of early preeclampsia and FGR a

(Cnossen et  al. 2008; Bahado-Singh and Jodicke 2010; O’Gorman et al. 2016a). >>While the negative predictive value is excellent in the presence of an  unremarkable Doppler (Bahado-Singh and Jodicke 2010), the sensitivity and positive predictive value is not suitable for use in general screening as a single marker, which is why combination with other markers is recommended (GabbayBenziv et  al. 2016; O’Gorman et  al. 2016a, b, 2017a, b; Yücel et al. 2016). >>With increasing uteroplacental perfusion disturbance, the fetoplacental compartment is also compromised.

The first sign is abnormalities in the flow pattern of the umbilical arteries (Baschat et  al. 2001). The assessment is quantitative (measurement of the resistance indices) on the one hand and qualitative with the assessment of the end-diastolic flow on the other. Absent end-diastolic flow occurs when approximately 60–70% of the villous vascular tree is defective (Vergani et al. 2005); further deterioration may result in reversed end-­diastolic flow in the umbilical arteries. b

UA LE

..      Fig. 9.25  a, b Doppler sonography of the uterine arteries: normal a and pathological b flow pattern of the uterine artery. UA LE left uterine artery

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a

b

c

d

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..      Fig. 9.26  a–d Doppler sonography of the umbilical arteries. Normal flow pattern of the umbilical artery a Doppler sonographic changes in the blood flow of the umbilical artery during fetal growth

restriction: b decreased end-diastolic flow and increased pulsatility index; c absent end-diastolic flow (zero-flow); d reversed end-diastolic flow.

Placental Function

Functional MRI examination offers the possibility to study vascularization, oxygenation and metabolism by means of different enhancement processes (Javor et  al. 2013; Siauve et  al. 2015; Mourier et  al. 2017). However, since contrast imaging using gadolinium is contraindicated, alternative contrast agents must be used (Siauve et al. 2015).

Assessment of placental function as the primary source of nutrition for the developing fetus is of paramount importance. In the past, various parameters (weights, measures and ratios) have been developed to assess placental function. However, all these parameters reflect the function of the placenta only very unsatisfactorily. In contrast, the pathological-­anatomical examination of the placenta, umbilical cord and fetal membranes is more informative (Graf 2008). Recent studies use magnetic resonance imaging (MRI) methods to investigate oxygenation and placental metabolism—in addition to blood flow assessment.

Placental Blood Flow Evaluation of placental blood flow by (three-­dimensional) power Doppler ultrasonography has been proposed as a potentially useful tool for predicting pregnancy complications (Odeh et  al. 2011ab). Inconsistent results and especially the low reproducibility

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(Cheong et  al. 2010) are reasons why this method has not found its way into routine diagnostics. Moore et  al. (2000a) reported indirect measurement of placental blood flow using magnetic resonance imaging (MRI). The technique called IVIM (Intravoxel Incoherent Motion) was able to show decreased perfusion in FGR (Moore et  al. 2000b). Subsequently, placental perfusion was measured with different MRI sequences (IVIM and ASL, “arterial spin labeling”) and correlated with the resistance (pulsatility index, PI) measured by Doppler sonography in the uterine arteries (Derwig et  al. 2013): It was shown that placental perfusion measured by MRI correlates with uterine Doppler and is reduced in pregnancies with small-for-gestational-age infants.

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Oxygenation and Placental Metabolism Intact placental oxygenation is essential for fetal growth and development. Various MRI techniques have been used in recent studies to investigate placental oxygenation and fetal oxygenation (Sørensen et  al. 2013; Huen et  al. 2013). However, these methods have not yet progressed beyond the research stage. MRI techniques are also used to study placental metabolism (Denison et al. 2012). Further studies must show whether this will be of importance in the future.

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Arlicot C, Herve P, Simon E, Perrotin F (2012) Three-­ dimensional surface rendering of the chorionic placental plate: the “tire” sign for the diagnosis of a circumvallate placenta. J Ultrasound Med 31(2):340–341 Bardo D, Oto A (2008) Magnetic resonance imaging for evaluation of the fetus and the placenta. Am J Perinatol 25(9):591–599 Becker RH, Vonk R, Mende BC, Ragosch V, Entezami M (2001) The relevance of placental location at 20–23 gestational weeks for prediction of placenta previa at delivery: evaluation of 8650 cases. Ultrasound Obstet Gynecol 17(6):496–501 Blaicher W, Brugger PC, Mittermayer C, Schwindt J, Deutinger J, Bernaschek G, Prayer D (2006) Magnetic resonance imaging of the normal placenta. Eur J Radiol 57(2):256–260 Blouin D, Rioux C (2012) Routine third trimester control ultrasound examination for low-lying or marginal placentas diagnosed at mid-pregnancy: is this indicated? J Obstet Gynaecol Can 34(5):425– 428 Bonel HM, Stolz B, Diedrichsen L, Frei K, Saar B, Tutschek B, Raio L, Surbek D, Srivastav S, Nelle M, Slotboom J, Wiest R (2010) Diffusionweighted MR imaging of the placenta in fetuses with placental insufficiency. Radiology 257(3):810–819 Bowman ZS, Kennedy AM (2014) Sonographic appearance of the placenta. Curr Probl Diagn Radiol 43(6):356–373 Brown DL, DiSalvo DN, Frates MC, Davidson KM, Genest DR (2002) Placental surface cysts detected on sonography: histologic and clinical correlation. J Ultrasound Med 21(6):641–646; quiz 647–648 Burton GJ, Jauniaux E, Charnock-Jones DS (2010a) The influence of the intrauterine environment on human placental development. Int J Dev Biol 54(2–3):303–312 Cooley SM, Donnelly JC, Walsh T, McMahon C, Gillen J, Geary MP (2011) The impact of ultrasonographic placental architecture on antenatal course, labor and delivery in a low-risk primigravid population. J Matern Fetal Neonatal Med 24(3):493–497 D’Antonio F, Iacovella C, Palacios-Jaraquemada J, Bruno CH, Manzoli L, Bhide A (2014) Prenatal identification of invasive placentation using magnetic resonance imaging: systematic review and meta-analysis. Ultrasound Obstet Gynceol 44(1):8–16 Dekan S, Linduska N (2011) Normal and pathological placental development: MRI and pathology. In: Prayer D (ed) Fetal MRI.  Springer, Berlin Heidelberg, pp 403–442 Dombrowski MP, Wolfe HM, Saleh A, Evans MI, O’Brien J (1992) The sonographically thick pla-

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alpha-­fetoprotein. Eur J Obstet Gynecol Reprod Biol 37(3):219–230 Jauniaux E, Ramsay B, Campbell S (1994) Ultrasonographic investigation of placental morphologic characteristics and size during the second trimester of pregnancy. Am J Obstet Gynecol 170(1 Pt 1):130–137 Jha P, Paroder V, Mar W, Horowtiz JM, Poder L (2016) Multimodality imaging of placental masses: a pictorial review. Abdom Radiol (NY) 41(12):2435–2444 Kalache KD, Espinoza J, Chaiworapongsa T, Londono J, Schoen ML, Treadwell MC, Lee W, Romero R (2003) Three-dimensional ultrasound fetal lung volume measurement: a systematic study comparing the multiplanar method with the rotational (VOCAL) technique. Ultrasound Obstet Gynecol 21(2):111–118 Kanne JP, Lalani TA, Fligner CL (2005) The placenta revisited: radiologic-pathologic correlation. Curr Probl Diagn Radiol 34(6):238–255 Kaplan CG (2008) Gross pathology of the placenta: weight, shape, size, colour. J Clin Pathol 61(12):1285–1295 Kofinas AD, Penry M, Swain M, Hatjis CG (1989) Effect of placental laterality on uterine artery resistance and development of preeclampsia and intrauterine growth retardation. Am J Obstet Gynecol 161(6 Pt 1):1536–1539 Krielessi V, Papantoniou N, Papageorgiou I, Chatzipapas I, Manios E, Zakopoulos N, Antsaklis A (2012) Placental pathology and blood pressure’s level in women with hypertensive disorders in pregnancy. Obstet Gynecol Int 2012:684083 Lax A, Prince MR, Mennitt KW, Schwebacjh JR, Budorick NE (2007) The value of specific MRI features in the evaluation of suspected placental invasion. Magn Res Imaging 25(1):87–93 Lee AJ, Bethune M, Hiscock RJ (2012) Placental thickness in the second trimester: a pilot study to determine the normal range. J Ultrasound Med 31(2):213–218 Linduska N, Dekan S, Messerschmidt A, Kasprian G, Brugger PC, Chalubinski K, Weber M, Prayer D (2009) Placental pathologies in fetal MRI and pathohistological correlation. Placenta 30(6):555– 559 Macara L, Kingdom JC, Kaufmann P, Kohnen G, Hair J, More IA, Lyall F, Greer IA (1996) Structural analysis of placental terminal villi from growth-restricted pregnancies with abnormal umbilical artery Doppler waveforms. Placenta 17(1):37–48 Masselli G, Brunelli R, Di Tola M, Anceschi M, Gualdi G (2011) MR imaging in the evaluation of placental abruption: correlation with sonographic findings. Radiology 259(1):222–230

McCarthy J, Thurmond AS, Jones MK, Sistrom C, Scanlan RM, Jacobson SL, Lowensohn R (1995) Circumvallate placenta: sonographic diagnosis. J Ultrasound Med 14(1):21–26 Meizner I, Mashiach R, Shalev Y, Ben-Rafael Z (1998) Blood flow velocimetry in the diagnosis of succenturiate placenta. J Clin Ultrasound 26(1):55 Merz E, Eichhorn KH, von Kaisenberg C, Schramm T, Arbeitsgruppe der DEGUM-Stufe III (2012) Updated quality requirements regarding secondary differentiated ultrasound examination in prenatal diagnostics (= DEGUM level II) in the period from 18 + 0 to 21 + 6 weeks of gestation. Ultraschall Med 33(6):593–596 Miwa I, Sase M, Torii M, Sanai H, Nahamura Y, Ueda K (2014) A thick placenta: a predictor of adverse pregnancy outcomes. Springerplus 3:353 Moldenhauer JS, Stanek J, Warshak C, Khoury J, Sibai B (2003) The frequency and severity of placental findings in women with preeclampsia are gestational age dependent. Am J Obstet Gynecol 189(4):1173–1177 Moran M, Ryan J, Higgins M, Brennan PC, McAuliffe FM (2011) Poor agreement between operators on grading of the placenta. J Obstet Gynaecol 31(1):24–28 Morikawa M, Cho K, Kataoka S, Kato EH, Yamada T, Yamada H, Minakami H (2005) Magnetic resonance image findings of placental lake: report of two cases. Prenat Diagn 25(3):250–252 Moscoso G, Jauniaux E, Hustin J (1991) Placental vascular anomaly with diffuse mesenchymal stem villous hyperplasia. A new clinico-pathological entity? Pathol Res Pract 187(2–3):324–328 Nyberg DA, Cyr DR, Mack LA, Wilson DA, Shuman WP (1987) Sonographic spectrum of placental abruption. AJR Am J Roentgenol 148(1):161–164 Odeh M, Ophir E, Maximovsky O, Grinin V, Bornstein J (2011a) Placental volume and three-dimensional power Doppler analysis in prediction of pre-­ eclampsia and small for gestational age between Week 11 and 13 weeks and 6 days of gestation. Prenat Diagn 31(4):367–371 Odibo AO, Goetzinger KR, Huster KM, Christiansen JK, Odibo L, Tuuli MG (2011) Placental volume and vascular flow assessed by 3D power Doppler and adverse pregnancy outcomes. Placenta 32(3):230–234 Oppenheimer L, Society of Obstetricians and Gynaecologists of Canada (2007) Diagnosis and management of placenta previa. J Obstet Gynaecol 29(3):261–273 Oppenheimer LW, Farine D, Ritchie JW, Lewinsky RM, Telford J, Fairbanks LA (1991) What is a low-lying placenta? Am J Obstet Gynecol 165(4 pt 1):1036–1038

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Oppenheimer L, Holmes P, Simpson N, Dabrowski A (2001) Diagnosis of low-lying placenta: can migration in the third trimester predict outcome? Ultrasound Obstet Gynecol 18(2):100–102 Oyelese Y, Smulian JC (2006) Placenta previa, placenta accreta, and vasa previa. Obstet Gynecol 107(4):927–941 Parveen Z, Tongson-Ignacio JE, Fraser CR, Killeen JL, Thompson KS (2007) Placental mesenchymal dysplasia. Arch Pathol Lab Med 131(1):131–137 Pereira N, Yao R, Guilfoil DS, Richard SD, Plante LA (2013) Placenta membranacea with placenta accrete: radiologic diagnosis and clinical implications. Prenat Diagn 33(13):1283–1286 Pomorski M, Zimmer M, Florjanski J, Michniewicz J, Wiatrowski A, Fuchs T, Milnerowicz-Nabzdyk E (2012) Comparative analysis of placental vasculature and placental volume in normal and IUGR pregnancies with the use of three-dimensional power Doppler. Arch Gynecol Obstet 285(2):331– 337 Porat S, Fitzgerald B, Wright E, Keating S, Kingdom JC (2013) Placental hyperinflation and the risk of adverse perinatal outcome. Ultrasound Obstet Gynecol 42(3):315–321 Proctor LK, Toal M, Keating S, Chitayat D, Okun N, Windrim RC, Smith GC, Kingdom JC (2009) Placental size and the prediction of severe earlyonset intrauterine growth restriction in women with low pregnancy-associated plasma proteinA. Ultrasound Obstet Gynecol 34(3):274–282 Proctor LK, Whittle WL, Keating S, Viero S, Kingdom JC (2010) Pathologic basis of echogenic cystic lesions in the human placenta: role of ultrasoundguided wire localization. Placenta 31(12):1111– 1115 Publications Committee, Society for Maternal-Fetal Medicine, Belfort MA (2010) Placenta accreta. Am J Obstet Gynecol 203(5):430–439 Raga F, Ballester MJ, Osborne NG, Bonilla-Musoles F (1996) Subchorionic placental cyst: a cause of fetal growth retardation–ultrasound and colorflow Doppler diagnosis and follow-up. J Natl Med Assoc 88(5):285–288 Raio L, Ghezzi F, Cromi A, Nelle M, Düring P, Schneider H (2004) The thick heterogeneous (jellylike) placenta: a strong predictor of adverse pregnancy outcome. Prenat Diagn 24(3):182–188 Reddy UM, Abuhamad AZ, Levine D, Saade GR (2014) Fetal imaging: executive summary of a joint. Eunice Kennedy Shriver National Institute of Child Health and Human Development, society for maternal-­ fetal medicine, American Institute of ultrasound in medicine, American College of Obstetricians and gynecologists, American College of radiology, society for pediatric radiology, and society of radiologists in ultra-

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sound fetal imaging workshop. J Ultrasound Med 33(5):745–757 Rizzo G, Capponi A, Pietrolucci ME, Aiello E, Arduini D (2012) First trimester placental volume and three dimensional power doppler ultrasonography in type I diabetic pregnancies. Prenat Diagn 32(5):480–484 Rosenberg T, Pariente G, Sergienko R, Wiznitzer A, Sheiner E (2011) Critical analysis of risk factors and outcome of placenta previa. Arch Gynecol Obstet 284(1):47–51 Sau A, Seed P, Langford K (2004) Intraobserver and interobserver variation in the sonographic grading of placental maturity. Ultrasound Obstet Gynecol 23(4):374–377 Schlembach D, Helmer H, Henrich W, von Heymann C, Kainer F, Korte W, Kühnert M, Lier H, Maul H, Rath W, Steppat S, Surbek D, Wacker J (2016) Peripartale Blutungen. Diagnostik und Therapie. Leilinie der Deutschen Gesellschaft für Gynäkologie und Geburtshilfe (S2k-Level, AWMF Register-Nr. 015/063, März 2016). http:// www.­awmf.­o rg/leitlinien/detail/ll/015-­0 63.­h tml. Accessed 16 Nov 2017 Schlensker KH (1971) Plazentographie mittels Ultraschallschnittbildverfahren. Geburtshilfe Frauenheilkd 31:879–897 Scott JS (1960) Placenta extrachorialis (placenta marginata and placenta circumvallata). J Obstet Gynaecol Br Emp 67:904–918 Seckin KD, Cakmak B, Karsli MF, Yeral MI, Gultekin IB, Oz M, Danisman N (2015) Is lateral localisation of placenta a risk factor for adverse perinatal outcomes? J Obstet Gynaecol 35(7):696–698 Silver RM, Landon MB, National Rouse DJ, Institute of Child Health and Human Development Maternal-­ Fetal Medicine Units Network et  al (2006) Maternal morbidity associated with multiple repeat cesarean deliveries. Obstet Gynecol 107(6):1226–1232 Soto E, Hernández-Andrade E (2015) Placenta, amniotic fluid, umbilical cord, and membranes. In: Kline-­Fath BM, Bulas DI, Bahado-Singh R (eds) Fundamental and advanced fetal imaging – ultrasound and MRI.  Wolters Kluver Health, Philadelphia, PA, pp 235–267 Souka AP, Papastefanou I, Michalitsi V, Papadopoulos GK, Kassanos D (2011) A predictive model of short cervix at 20–24 weeks using first-trimester cervical length measurement and maternal history. Prenat Diagn 31(2):202–206 Suzuki S (2008) Clinical significance of pregnancies with circumvallate placenta. J Obstet Gynaecol Res 34(1):51–54 Suzuki S, Igarashi M (2008) Clinical significance of pregnancies with succenturiate lobes of placenta. Arch Gynecol Obstet 277(4):299–301

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Thompson MO, Vines SK, Aquilina J, Wathen NC, Harrington K (2002) Are placental lakes of any clinical significance? Placenta 23(8–9):685–690 Tikkanen M, Nuutila M, Kiilesmaa V, Paavonen J, Ylikorkala O (2006) Clinical presentation and risk factors of placental abruption. Acta Obstet Gynecol Scand 85(6):700–705 Todros T, Sciarrone A, Piccoli E, Guiot C, Kaufmann P, Kingdom JC (1999a) Umbilical Doppler waveforms and placental villous angiogenesis in pregnancies complicated by fetal growth restriction. Obstet Gynecol 93(4):499–503 Viero S, Chaddha V, Alkazaleh F, Simchen MJ, Malik A, Kelly E, Windrim R, Kingdom JC (2004) Prognostic value of placental ultrasound in pregnancies complicated by absent end-diastolic flow velocity in the umbilical arteries. Placenta 25(8– 9):735–741 Vinnars MT, Nasiell J, Ghazi S, Westgren M, Papadogiannakis N (2011) The severity of clinical manifestations in preeclampsia correlates with the amount of placental infarction. Acta Obstet Gynecol Scand 90(1):19–25 Walker MG, Fitzgerald B, Keating S, Ray JG, Windrim R, Kingdom JC (2012) Sex-specific basis of severe placental dysfunction leading to extreme preterm delivery. Placenta 33(7):568–571 Yampolsky M, Salafia CM, Shlakhter O, Haas D, Eucker B, Thorp J (2008) Modeling the variability of shapes of a human placenta. Placenta 29(9):790–797 Yang Q, Wen SW, Oppenheimer L, Chen XK, Black D, Gao J, Walker MC (2007) Association of caesarean delivery for first birth with placenta praevia and placental abruption in second pregnancy. BJOG 114(5):609–613

Section 9.2 Bahado-Singh RO, Jodicke C (2010) Uterine artery Doppler in first-trimester pregnancy screening. Clin Obstet Gynecol 53(4):879–887 Baron J, Shwarzman P, Sheiner E, Weintraub AY, Spiegel E, Sciaky Y, Dukler D, Hershkovitz R (2015) Blood flow Doppler velocimetry measured during active labor. Arch Gynecol Obstet 291(4):837–840 Baschat AA, Gembruch U, Harman CR (2001) The sequence of changes in Doppler and biophysical parameters as severe fetal growth restriction worsens. Ultrasound Obstet Gynecol 18(6):571–577 Baschat AA, Cosmi E, Bilardo CM, Wolf H, Berg C, Rigano S, Germer U, Moyano D, Turan S, Hartung J, Bhide A, Müller T, Bower S, Nicolaides KH, Thilaganathan B, Gembruch U, Ferrazzi E, Hecher K, Galan HL, Harman CR (2007) Predictors of neonatal outcome in early-onset

placental dysfunction. Obstet Gynecol 109(2 Pt 1):253–261 Burton GJ, Fowden A (2015) The placenta: a multifaceted, transient organ. Phil Trans R Soc 370(1663):20140066 Burton GJ, Watson AL, Hempstock J, Skepper JN, Jauniaux E (2002) Uterine glands provide histiotrophic nutrition for the human fetus during the first trimester of pregnancy. J Clin Endocrinol Metab 87(6):2954–2959 Burton GJ, Jauniaux E, Charnock-Jones DS (2010b) The influence of the intrauterine environment on human placental development. Int J Dev Biol 54(2–3):303–312 Chaoui R, Heling KS, Karl K (2014) Ultrasound of the fetal veins part 1: the intrahepatic venous system. Ultraschall Med 35(3):208–228 Cheong KB, Leung KY, Li TK, Chen HY, Lee YP, Tang MH (2010) Comparison of inter- and intraobserver agreement and reliability between three different types of placental volume measurement technique (XI VOCAL, VOCAL and multiplanar) and validity in the in-vitro setting. Ultrasound Obstet Gynecol 36(2):210–217 Cnossen JS, Morris RK, ter Riet G, Mol BW, Post JA van der, Coomarasamy A, Zwinderman AH, Robson SC, Bindels PJ, Kleijnen J, Khan KS (2008) Use of uterine artery Doppler ultrasonography to predict pre-­eclampsia and intrauterine growth restriction: a systematic review and bivariable meta-analysis. CMAJ 178(6):701–711 Denison FC, Semple SI, Stock SJ, Walker J, Marshall I, Norman JE (2012) Novel use of proton magnetic resonance spectroscopy (1HMRS) to non-invasive assess placental metabolism. PLoS One 7(8):e42926 Derwig I, Barker GJ, Poon L, Zelaya F, Gowland P, Lythgoe DJ, Nicolides K (2013) Association of placental T2 relaxation times and uterine artery Doppler ultrasound measures of placental blood flow. Placenta 34(6):474–479 Everett TR, Lees CC (2012) Beyond the placental bed: placental and systemic determinants of the uterine artery waveform. Placenta 33:893–901 Ferrazzi E, Bozzo M, Rigano S, Bellotti M, Morabito A, Pardi G, Battaglia FC, Galan HL (2002) Temporal sequence of abnormal Doppler changes in the peripheral and central circulatory systems of the severely growth-restricted fetus. Ultrasound Obstet Gynecol 19(2):140–146 Gabbay-Benziv R, Oliveira N, Baschat AA (2016) Optimal first trimester preeclampsia prediction: a comparison of multimarker algorithm, risk profiles and their sequential application. Prenat Diagn 36(1):34–39 Giles WB, Trudinger BJ, Baird PJ (1985) Fetal umbilical artery flow velocity waveforms and placental

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resistance: pathological correlation. Br J Obstet Gynaecol 92(1):31–38 Gómez O, Figueras F, Fernández S, Bennasar M, Martínez JM, Puerto B, Gratacós E (2008) Reference ranges for uterine artery mean pulsatility index at 11–41 weeks of gestation. Ultrasound Obstet Gynecol 32(2):128–132 Graf AH (2008) Morphologie, Physiologie und Pathologie des maternoplazentaren, fetoplazentaren und fetalen Kreislaufs. In: Steiner H, Schneider KTM (eds) Dopplersonographie in Geburtshilfe und Gynäkologie. Springer Medizin, Heidlberg, pp 3–11 Hernandez-Andrade E, Brodszki J, Lingman G, Gudmundsson S, Molin J, Marsál K (2002) Uterine artery score and perinatal outcome. Ultrasound Obstet Gynecol 19(5):438–442 Hitschold T, Weiss E, Beck T, Hünterfering H, Berle P (1993) Low target birth weight or growth retardation? Umbilical Doppler flow velocity waveforms and histometric analysis of fetoplacental vascular tree. Am J Obstet Gynecol 168(4):1260–1264 Huen I, Morris DM, Wright C, Parker GJ, Sibley CP, Johnstone ED, Naish JH (2013) R1 and R2 * changes in the human placenta in response to maternal oxygen challenge. Magn Reson Med 70(5):1427–1433 Jauniaux E, Watson AL, Hempstock J, Bao YP, Skepper JN, Burton GJ (2000) Onset of maternal arterial blood flow and placental oxidative stress. A possible factor in human early pregnancy failure. Am J Pathol 157(6):2111–2122 Javor D, Nasel C, Schweim T, Dekan S, Chalubinski K, Prayer D (2013) In vivo assessment of putative functional placental tissue volume in placental Intrauterine Growth Restriction (IUGR) in human fetuses using diffusion tensor magnetic resonance imaging. Placenta 34(8):676–680 Kehl S, Dötsch J, Gembruch U, Hecher K, Schmitz D, Schlembach D, Stepan H (2017) Intrauterine growth restriction. Guideline of the German Society of Gynecology and Obstetrics (S2k, AWMF-­ Registry-­ No.: 015/080, October 2016). http://www.­awmf.­org/leitlinien/detail/ll/015-­080.­ html. Accessed 13 Nov 2017 Krebs C, Macara LM, Leiser R, Bowman AW, Greer IA, Kingdom JC (1996) Intrauterine growth restriction with absent end-diastolic flow velocity in the umbilical artery is associated with maldevelopment of the placental terminal villous tree. Am J Obstet Gynecol 175(6):1534–1542 Messerschmidt A, Baschat A, Linduska N, Kasprian G, Brugger PC, Bauer A, Weber M, Prayer D (2011) Magnetic resonance imaging of the placenta identifies placental vascular abnormalities independently of Doppler ultrasound. Ultrasound Obstet Gynecol 37(6):717–722

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Monaghan C, Binder J, Thilaganathan B, Morales-­ Roselló J, Khalil A (2018) Perinatal loss at term: the role of uteroplacental and fetal Doppler assessment. Ultrasound Obstet Gynecol. 52(1):72–77. https://doi.org/10.1002/uog.17500 Moore RJ, Issa B, Tokarczuk P, Duncan KR, Boulby P, Baker PN, Bowtell RW, Worthington BS, Johnson IR, Gowland PA (2000a) In vivo intravoxel incoherent motion measurement in the human placenta using echo-planar imaging at 0.5 T. Magn Reson Med 43(2):295–302 Moore RJ, Strachan BK, Tyler DJ, Duncan KR, Baker PN, Worthington BS, Johnson IR, Gowland PA (2000b) In utero perfusing fraction maps in normal and growth restricted pregnancy measured using IVIM echo-planar MRI.  Placenta 21(7):726–732 Mourier E, Tarrade A, Duan J, Richard C, Bertholdt C, Beaumont M, Morel O, Chavatte-Palmer P (2017) Non-invasive evaluation of placental blood flow: lessons from animal models. Reproduction 153(3):R85–R96 Myatt L (1992) Control of vascular resistance in the human placenta. Placenta 13(4):329–341 O’Gorman N, Tampakoudis G, Wright A, Wright D, Nicolaides KH (2016a) Uterine artery pulsatility index at 12, 22, 32 and 36 weeks’ gestation in screening for pre-eclampsia. Ultrasound Obstet Gynecol 47(5):565–572 O’Gorman N, Wright D, Syngelaki A, Akolekar R, Wright A, Poon LC, Nicolaides KH (2016b) Competing risks model in screening for preeclampsia by maternal factors and biomarkers at 11–13 weeks gestation. Am J Obstet Gynecol 214(1):103.e1–103.e12 O’Gorman N, Wright D, Poon LC, Rolnik DL, Syngelaki A, de Alvarado M, Carbone IF, Dutemeyer V, Fiolna M, Frick A, Karagiotis N, Mastrodima S, de Paco MC, Papaioannou G, Pazos A, Plasencia W, Nicolaides KH (2017a) Multicenter screening for pre-­eclampsia by maternal factors and biomarkers at 11–13 weeks’ gestation: comparison with NICE guidelines and ACOG recommendations. Ultrasound Obstet Gynecol 49(6):756–760 O’Gorman N, Wright D, Poon LC, Rolnik DL, Syngelaki A, Wright A, Akolekar R, Cicero S, Janga D, Jani J, Molina FS, de Paco MC, Papantoniou N, Persico N, Plasencia W, Singh M, Nicolaides KH (2017b) Accuracy of competingrisks model in screening for pre-­ eclampsia by maternal factors and biomarkers at 11–13 weeks’ gestation. Ultrasound Obstet Gynecol 49(6):751– 755 Odeh M, Ophir E, Maximovsky O, Grinin Bornstein J (2011b) Placental volume and three-dimensional power Doppler analysis in prediction of pre-­

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eclampsia and small for gestational age between Week 11 and 13 weeks and 6 days of gestation. Prenat Diagn 31(4):367–371 Osol G, Moore LG (2014) Maternal uterine vascular remodeling during pregnancy. Microcirculation 21(1):38–47 Pasca AM, Penn AA (2010) The placenta: the lost neuroendocrine organ. Neoreviews 11(2):e64–e77 Redman CW, Sargent IL (2005) Latest advances in understanding preeclampsia. Science 308(5728):1592–1594 Sheppard BL, Bonnar JJ (1981) An ultrastructural study of utero-placental spiral arteries in hypertensive and normotensive pregnancy and fetal growth retardation. Br J Obstet Gynaecol 88(7):695–705 Shilling C, Walsh C, Downey P, Mooney E (2014) Umbilical artery thrombosis is a rare but clinically important finding: a series of 7 cases with clinical outcomes. Pediatr Dev Pathol 17(2):89–93 Siauve N, Chalouhi GE, Deloison B, Alison M, Clement O, Ville Y, Salomon LJ (2015) Functional imaging of the human placenta with magnetic resonance. Am J Obstet Gynecol 213(4):103–114 Sørensen A, Peters D, Fründ E, Lingman G, Christiansen O, Uldbjerg N (2013) Changes in

human placental oxygenation during maternal hyperoxia estimated by blood oxygen level-dependent magnetic resonance imaging (BOLD MRI). Ultrasound Obstet Gynecol 42(3):310–314 Todros T, Sciarrone A, Piccoli E, Guiot C, Kaufmann P, Kingdom J (1999b) Umbilical Doppler waveforms and placental villous angiogenesis in pregnancies complicated by fetal growth restriction. Obstet Gynecol 93(4):499–503 Vergani P, Roncaglia N, Locatelli A, Andreotti C, Crippa I, Pezzullo JC, Ghidini A (2005) Antenatal predictors of neonatal outcome in fetal growth restriction with absent end-diastolic flow in the umbilical artery. Am J Obstet Gynecol 193(3 Pt 2):1212–1218 Voigt HJ, Becker V (1992) Doppler flow measurements and histomorphology of the placental bed in uteroplacental insufficiency. J Perinat Med 20(2):139–147 Yücel B, Gedikbasi A, Dündar O, Olgac Y, Yildirim D, Yildirim G, Polat I (2016) The utility of first trimester uterine artery Doppler, placental volume and PAPP-A levels alone and in combination to predict preeclampsia. Pregnancy Hypertens 6(4):269–273

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Disorders of Early Pregnancy and Pregnancy Loss Stephanie Pildner von Steinburg, Ekkehard Schleußner, Ruben Kuon, Kilian Vomstein, and Bettina Toth Contents 10.1

Early Pregnancy and Its Disturbance – 220

10.1.1 10.1.2

 iagnosis of Early Pregnancy – 220 D Pregnancy Loss – 224

10.2

Recurrent Pregnancy Loss – 229

10.2.1 10.2.2 10.2.3

I ntroduction – 229 Established Risk Factors – 230 Possible New Risk Factors – 234

References – 238

© Springer-Verlag GmbH Germany, part of Springer Nature 2023 B. Huppertz, E. Schleußner (eds.), The Placenta, https://doi.org/10.1007/978-3-662-66256-4_10

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10.1 

 arly Pregnancy and Its E Disturbance

Stephanie Pildner von Steinburg and Ekkehard Schleußner

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First published in Schneider et  al. (eds.) (2016) The obstetrician. 5th edn., p.  21 slightly modified. Clinical, laboratory chemistry and sonographic methods allow early and reliable diagnosis and assessment of early pregnancy. Serial determination of human chorionic gonadotropin (hCG)—taking into account appropriate doubling times—allows differentiation between intact and disturbed implantation. Transvaginal sonography can provide information on the site of implantation, the vitality of the embryo and the presence of a multiple pregnancy. In addition, sonographic assessment allows a largely reliable determination of the gestational age. The term “pregnancy loss” is understood to stand for a (non-artificial) loss of pregnancy before the child becomes viable. It is of importance to distinguish between sporadic and recurrent pregnancy losses, whereby a recurrent pregnancy loss occurs when there are three or more consecutive miscarriages. In the following, the clinical stages and the various causes of sporadic pregnancy losses will be discussed.

10.1.1 

Diagnosis of Early Pregnancy

General Remarks The absence of regular menstruation is the most common indication that pregnancy has occurred, and a detailed history of the menstrual cycle can support this suspicion. Subjective complaints of early pregnancy include symptoms such as the unusual rejection of certain stimulants and foods, morning sickness with vomiting, breast ten-

derness and emotional imbalance. Also very early in pregnancy, symptoms such as increased vaginal discharge and a tendency to constipation and pollakiuria may appear as indications of altered bowel and bladder function. These uncertain signs of pregnancy (which by definition also include absence of menorrhea) can appear in varying degrees. On gynecological examination, further uncertain indications of pregnancy include loosening and livid discoloration of vulva, introitus, vagina and cervix. The loosening of the uterus and in particular of the lower uterine segment, whereat the fingers of the inner and outer hand can almost touch each other on palpation (Hegar pregnancy sign), has been evaluated in the past as a valuable indication of a possibly existing early pregnancy. Often, the enlarged ovary can be palpated with the corpus luteum graviditatis already in the early stage of pregnancy. A palpable enlargement of the uterus, on the other hand, is found at the earliest from the 7th–8th week of pregnancy, but the interindividual variability is known to be high. However, the clinical findings provide only limited information about the presence, integrity and regular growth of an intrauterine early pregnancy, which is why the use of laboratory chemistry and sonographic methods appears to be useful in answering these questions.

Laboratory Diagnostics Human Chorionic Gonadotropin (hCG) The glycoprotein “human chorionic gonadotropin” (hCG) consists of two subunits (α- and β-chain) and is expressed in trophoblast cells. hCG is detectable in maternal serum at the earliest 8 days after ovulation and shows similarities to the luteinizing hormone (LH) with regard to molecular structure and luteotrophic effect. The different structure of the β-chains allows—with the aid of specific antibodies—the differentiation between (β-)hCG and LH in the common

221 Disorders of Early Pregnancy and Pregnancy Loss

test systems. When pregnancy occurs, hCG increasingly replaces LH, whereat the maintenance of steroid synthesis in the corpus luteum is in the foreground. Here, a sufficiently high progesterone level in the serum is important for the maintenance of early pregnancy. In the first 10–12 days of an intact intrauterine singleton pregnancy, the doubling time of hCG in serum is about 1.3 days. With increasing gestational age and/or higher hCG levels, the doubling time is prolonged—for example, it is reported to be 3 days at levels between 1200 and 6000 mIE/ ml. At this point, however, it should be emphasized that lower initial values and/or a prolonged doubling time of hCG should not be taken as the sole indication of extrauterine pregnancy or disturbed intrauterine early pregnancy. Conversely, short doubling times and/or high levels of hCG do not necessarily indicate the presence of trophoblastic disease. >>The highest hCG levels with 50,000– 100,000 mIE/ml are found in the tenth week of gestation, after which there is a continuous drop to values around 10,000–20,000 mIE/ml up to the 20th week of gestation. These values remain more or less constant until delivery of the child (Speroff et al. 1994).

Additional Factors, Hormones and Screening Tests Progesterone is secreted in a pulsatile manner, therefore the values fluctuate between two and 40 ng/ml within a very short time. Due to this, they do not play a major role in practice for the assessment of the regular development of a pregnancy; however, values >An intact early pregnancy is only confirmed by evidence of embryonic cardiac action.

Since the heart does not begin to pulsate until the 23rd embryonic day post conception, it cannot be visualized on ultrasound before 5 + 2 weeks gestation. At the latest from the seventh week of gestation, corresponding to a CRL of 6–9 mm, it must be possible to show a clear cardiac action. In the course of early pregnancy, the heart rate initially increases to 180 beats per minute (bpm) up to the ninth/tenth week of gestation, and then falls to about 140 bpm at the end of the first trimester. In early ultrasound, the placenta cannot be localized because physiologically it lines the amniotic cavity in a circular fashion as a chorion rich in echoes (. Figs.  10.1 and 10.3). It is not until the end of the first trimester that the majority of the chorion regresses and forms the chorion laeve, and then later fuses with the amniotic membrane to form the fetal membranes. In the area of the umbilical cord insertion, the fetal vessels will sprout into the developing placenta and then form the differentiated placenta in this area, which can then also be delineated sonographically. At this early stage, on the other hand, the detection of a multiple pregnancy is unproblematic. The detection of chorionicity and  

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0

..      Fig. 10.3  a Embryo in the eighth week of gestation, CRL 29 mm, arm and leg buds can be displayed on both sides. b Growth curves of the crown-rump length from the German maternity record

a

b

c

d

..      Fig. 10.4  a–c Differentiation of chorionicity and amnionicity in first trimester multiples. a Dichorialdiamnial (2 chorionic sacs, 2 yolk sacs, 2 embryos); b

monochorial-diamnial (1 chorionic sac, 2 yolk sacs, 2 embryos); c monochorial-monoamnial (1 chorionic sac, 1 yolk sac, 2 embryos); d normal singleton pregnancy

amnioticity is also much easier than in later weeks of pregnancy, but of crucial importance for the management of possible complications of multiple pregnancy. This is why this must also be documented in the maternity record at the first ultrasound screening. Sonographic indicators of multiple pregnancy are the number of chorionic cavities, yolk sacs, and embryos (. Fig. 10.4). The lambda sign, which occurs in a dichorial situation and represents placental tissue between the two amniotic membranes,

also serves to differentiate mono- and dichorial twin pregnancies (. Fig. 10.5). Already in the first trimester of pregnancy, there are indications for the presence of fetal diseases (conspicuous body contour with regard to neck and abdominal wall, abnormal heart rate, abnormal organ structures, abnormalities of the umbilical cord and/or placenta, conspicuous growth curve, etc.), which can be detected in the first trimester screening in the sense of an early diagnosis of malformation (Merz et al. 2004).

 

 

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can lead to miscarriages in the first and second trimesters are dealt with together in the corresponding sections of the chapter. So far, little attention has been paid to the clinical significance that the history of miscarriages has for the outcome of an existing pregnancy: In addition to the increasing probability of another miscarriage, the risk of a (very early) preterm birth, caused by premature rupture of the membranes or preterm labor, doubles already after the first miscarriage (Buchmayer et al. 2004).

View

..      Fig. 10.5  Lambda sign in dichorial-diamnial twins

Clinically, a threatened abortion (abortus imminens) is accompanied by vaginal bleeding with or without uterine contractions, Definition and Epidemiology usually without opening of the external cerThe term “pregnancy loss” refers to a (non-­ vical os. Sonographically, a vital embryo/ artificial) loss of pregnancy before the child fetus, a preserved uterine cervix and a closed is viable, defined by a birth weight of 50%. 2010). There are also no satisfactory conSporadic miscarriages are distinguished trolled studies on drug treatment with profrom recurrent miscarriages, which are defined gestogens and/or hCG.  However, there as three or more consecutive miscarriages. seems to be evidence that the use of progesFrom a clinical point of view, a division into terone orally (50–100 mg per day), dydrogesearly pregnancy losses up to the 12th–14th terone (10  mg two times per day) or hCG week of gestation and late pregnancy losses (5000 I.U. weekly to three times 9000 I.U. from the 14th week of gestation onwards weekly) can reduce the rate of miscarriages makes sense. However, with regard to the eti- later in life (meta-analyses in Devaseelan ology, except for endocrine causes and cervi- et al. 2010; Wahabi et al. 2011; Carp 2012, cal insufficiency, the transition between early 2015). However, the high rate of chromoand late pregnancy losses seems to be smooth. somal anomalies in sporadic miscarriages For this reason, the etiological factors that (see below) must be taken into account, and 10.1.2 

10

 linical Stages of the Abortion C Process

Pregnancy Loss

225 Disorders of Early Pregnancy and Pregnancy Loss

none of the therapeutic measures indicated can change the outcome of the pregnancy. If cardiac actions are present in the first trimester when abortion is threatened, a favorable course of pregnancy can be expected in about 90% of cases. This is of high practical importance for the reassurance of the affected patient. In incipient abortion (abortus incipiens) there is a softening and shortening of the uterine cervix with dilatation of the cervical canal under labor-like uterine contractions. In this situation, sonography allows confirmation of the clinical findings at the cervix—in addition, a deformed chorionic cavity with a non-vital embryo/fetus and a pronounced perichorial hematoma are often found (. Fig. 10.6). If partial or complete spontaneous expulsion of the embryo/fetus and the placenta has already occurred, an incomplete or complete abortion is present. On clinical examination, the uterus is often smaller than is appropriate for the gestational age, and a gaping cervical canal is found, although this may have reformed in the case of a complete abortion. Sonography cannot detect an intrauterine chorionic cavity with a vital embryo. Difficulties are often caused by the sonographic differentiation between placental remnants remaining in utero and blood clots, each of which may appear as irregular  

10

echo-dense structures. Since both clinically and sonographically a differentiation between a complete and incomplete abortion is not possible with certainty, a vacuum curettage can be offered—especially in cases of persistent bleeding. In a missed abortion there is no spontaneous abortion despite the fact that the embryo/fetus has died or is no longer developing. Clinically, the uterus is smaller than corresponding to the period of amenorrhea, the cervix appears coarse, and the cervical canal is closed. In the context of hCG determinations, it is not a possibly lowered baseline value that is of importance, but the absence or delayed doubling of the values within a period of time. On ultrasound, a non-vital embryo/fetus is found passively following the movements of the amniotic fluid in the course of a push palpation of the uterus. From a CRL of 7 mm, cardiac actions must be sonographically detectable. Biometry often shows an embryo that is too small in relation to the chorionic cavity diameter and to the calculated gestational age. If no embryonic parts can be visualized with a mean chorionic diameter > 25 mm, a “blighted ovum” must be assumed. In cases of prolonged missed abortion, severe coagulation disorders with the development of disseminated intravascular coagulopathy (DIC) have been reported in

Hematoma

..      Fig. 10.6  Amorphous embryo and perichorial hematoma in missed abortion

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individual cases. However, this is a very rare complication. The rule used to be that curettage was necessary for retained pregnancy material after abortion to prevent infection. However, a three-arm study comparing expectative with medical (vaginal misoprostol) and surgical procedures in 1200 women with missed abortion or incomplete abortion before 13 completed weeks of gestation (Trinder et al. 2006) showed no differences in (low) infection rates between the two collectives. The rates of post curettage (indicated because of heavy bleeding or sonographic suspicion of retention after 14 days) were 44% vs. 13% vs. 5%. For incomplete abortion, prospective management showed a success rate of 75%, but with a rate of unplanned inpatient admissions of 29%. For missed abortion, the proportion of women requiring surgical intervention was lowest in the drug treatment arm at 38%, but with longer hospital stays. A recent Cochrane analysis (Kim et  al. 2017) also concluded that medication or wait-and-see is an acceptable alternative to curettage based on outcome, but the data for fertility preservation of the different methods is not yet robust. If the cervix is closed, prostaglandins should be used preoperatively to soften the cervix in order to prevent surgical complications and subsequent cervical insufficiencies. Gemeprost or misoprostol (off-label use!) are available for this purpose. In surgical intervention, suction curettage should be preferred because of the lower complication rates. If increased bleeding occurs during/after surgical uterine evacuation, drug-based augmentation of uterine contractions must be performed as in the case of atonic postpartum hemorrhage. If the size of the gestational sac corresponds to that of the second trimester, a medically assisted spontaneous expulsion should first be attempted. Here, a subsequent curettage is usually necessary to

remove any remaining placental remnants. Various prostaglandins (Gemeprost, Sulproston, Dinoproston, Minprostin and Misoprostol) as well as Oxytocin are used depending on the maturity of the cervix. With regard to perioperative antibiotic prophylaxis, various substances, dosages and administration methods have been tested: The optimal regimen could not yet be determined on an evidence level due to the many different therapeutic approaches, but tetracyclines, ceftriaxone and metronidazole can reduce post-interventional infection rates with manageable side effects (overview in Morrill et al. 2013). The presence of an infected abortion is indicated by an increase in temperature > 38 °C, a leukocytosis of ≥12,000 and a significant increase in the erythrocyte sedimentation rate and C-reactive protein (CRP) level. In most cases, the clinical stage of an incipient or incomplete abortion is present, and in this situation, a previous attempted abortion must also be considered. In the initial stage, the infection affects the uterine cavity, but in the further course it may spread to the parametria with adnexa, to the peritoneum and hematogenously in the sense of a septic abortion. A frequently underestimated early symptom of an incipient septic event is persistent hypotension with tachycardia, which may initially be misinterpreted as vegetative dystonia or as a consequence of volume deficiency. In the further course, septic shock may develop with the cardinal symptoms of circulatory and renal failure as well as manifest disseminated intravascular coagulopathy (DIC) with a mortality rate  of about 20% (Finkielman et al. 2004). The specific therapy of an infected or septic abortion should therefore be started as early as possible and comprises, after obtaining bacterial cultures, 55 high-dose intravenous antibiotics (e.g. clindamycin + gentamycin ± ampicillin or ampicillin + gentamycin + metronidazole or levofloxacin + metronidazole or

10

227 Disorders of Early Pregnancy and Pregnancy Loss

imipenem or single substances with a similar spectrum), 55 adequate volume substitution and 55 the control of coagulation parameters, in order to be able to detect an incipient DIC as early as possible. In the case of a uterine size >14 weeks of gestation, a drug-assisted expulsion can be aimed at the same time. Before the 14th week of gestation, surgical emptying or palpation of the uterus can be planned immediately after the start of adequately dosed intravenous antibiotic therapy, as the risk of further spread of septicemia in connection with the surgical intervention is low. Due to the antibiotics available nowadays, this procedure has proven to be more effective than waiting too long and the risk of developing endotoxin shock.

 auses of Pregnancy Loss C Chromosomal Abnormalities in Pregnancy Tissue Numerical chromosomal abnormalities are detected in 50–70% of all sporadic miscarriages, in 90% of early miscarriages, in 50% of miscarriages in the 8th–11th week of gestation, 30% in the 16th–19th week of gestation and 6–12% beyond the 20th week of gestation (overview in Warren and Silver 2008). The most common of these aneuploidies are autosomal trisomies (60%) (most commonly trisomy 16, 20–30% of trisomies) and monosomy X (20%) and polyploidies (20%) followed by structural anomalies (Warren and Silver 2008). Mosaics confined to the chorion or placenta are also found more frequently in sporadic pregnancy losses (Kalousek et  al. 1992). In contrast, abnormalities in the embryonic/fetal k ­ aryotype are not among the common causes of recurrent pregnancy losses. On the contrary, a normal set of chromosomes is often found in the pregnancy tissue of couples with recurrent pregnancy losses (Sullivan et al. 2004).

Genetic Causes in the Parents Genetic causes of recurrent pregnancy losses include chromosomal abnormalities of one parent, molecular defects, and multifactorial syndromes. Mutations and genetic defects may be responsible for a significant proportion of euploid pregnancy losses. However, the molecular genetic techniques to detect more of such associations have only recently become available, and systematic insights into the role of mutations in causing recurrent pregnancy losses are still lacking.

Uterine Anomalies Congenital Uterine Anomalies There is no reliable information on the general incidence of congenital uterine anomalies. In women with recurrent pregnancy loss, uterine malformations are reported in  10–30%  of cases, while the risk of pregnancy loss is likely to depend primarily on the type of anomaly present and its severity. In some cases, the literature contains very different data on the rate of pregnancy losses in the various uterine malformations (. Table 10.1). In general, the increased risk of obstetric complications in all malformations of the Müllerian ducts should be mentioned, such as prematurity, fetal growth restriction, anomalies of the position and pole onfigurations and uterine ruptures after surgical correction.  

..      Table 10.1  Risk of pregnancy loss depending on the type of uterine malformation. (Overview in Grimbizis et al. 2001) Risk of pregnancy loss

Frequency (%)

Uterus septus/subseptus

44.3

Uterus unicornis

36.5

Uterus bicornis

36

Uterus didelphys

32.2

Uterus arcuatus

25.7

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Acquired Uterine Anomalies

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Intrauterine synechiae after endometritis and intrauterine surgery and, more recently, after transcavitary compression sutures, described in the context of atony treatment after a previous delivery, are also considered a risk factor for early and late pregnancy losses as well as for placental disorders (Poujade et al. 2011). The incidence of intrauterine synechiae and their extent increases with the number of previous losses and intrauterine interventions: for example, the incidence of such adhesions is 14–16% after two, but 32% after three or more early losses (Friedler et al. 1993). Even large myoma often remain asymptomatic during pregnancy. However, the presence of submucosal myoma is associated with an increased risk of preterm and late losses and other pregnancy complications (preterm births, premature placental abruption). The importance of cervical insufficiency in triggering pregnancy losses—mostly in the second trimester—has probably been overestimated in the past and is likely to be around 1% in unselected pregnant women, but around 13% in pregnant women with a history of recurrent pregnancy losses (Stray-­ Pedersen and Stray-Pedersen 1984). The recurrence risk of late pregnancy loss or extreme preterm birth in cervical insufficiency is 28% (Sneider et al. 2016). The clinical presentation of cervical insufficiency may be gradual, painless dilatation of the cervix, bulging of the amnion, or premature rupture of the membranes with a uterus without labor. However, any of these symptoms may also occur during the course of a pregnancy loss of a different cause and therefore does not prove the causality of cervical insufficiency for the loss of pregnancy.

Infections A number of bacterial, parasitic and viral infections are causally associated with sporadic pregnancy loss. The associations of

syphilis, listeriosis and toxoplasmosis as well as various viral infections with pregnancy outcome are well known. Lyme disease is also still discussed as a cause of miscarriages and malformations. Bacterial vaginosis is a recognized risk factor for late pregnancy losses, preterm births and premature rupture of membranes. However, an association with pregnancy losses during the first trimester was not confirmed in a meta-analysis (van Oostrum et  al. 2013), while there was an association with loss of preclinical pregnancies. In addition, when assessing the causality of bacterial vaginosis for the occurrence of pregnancy losses, the coincidence with other genital infections (chlamydia, mycoplasma) that potentially cause miscarriages must also be taken into account.

Stimulants and Pollutants Nicotine and/or increased caffeine consumption seem to be associated with an increased risk of miscarriage—however, these associations have not been proven in all studies. Significantly increased numbers of miscarriages, on the other hand, are found in women with chronic alcohol, opiate and cocaine abuse during pregnancy. Occupational exposure to cytostatic drugs and anesthetic gases has been recorded to increase the rate of pregnancy losses among medical personnel. Workers in certain sectors of the metal industry, in chemical or pharmaceutical plants, in dry cleaning and women handling organic solvents or paints also appear to have an increased risk of miscarriage, although the differences with the miscarriage rates of unexposed women appear to be significant only for certain substances or combinations. In view of the large number of substances in question and the variable duration and intensity of exposure, the partly contradictory findings on the role of pollutants at the workplace and in the domestic environment in triggering pregnancy losses are not surprising, especially since knowledge of the

229 Disorders of Early Pregnancy and Pregnancy Loss

fertility-inhibiting potential of individual substances and their additive effects is still fragmentary.

Endocrine Causes Obesity has generally been shown to be a risk factor for miscarriage (Boots and Stephenson 2011), not only in the field of assisted reproduction or in the presence of polycystic ovary syndrome (PCOS). However, data demonstrating that normalizing body weight—in addition to regulating the menstrual cycle— can also reduce the risk of miscarriage are lacking to date (Best et al. 2017). A prediabetic metabolic state, recognizable by an elevated HOMA index (ratio of fasting blood glucose to insulin) or a pathological oral glucose tolerance test, can also cause recurrent pregnancy losses. Preconceptional therapy with metformin can be promising in this case. In the course of pregnancy, insulin therapy is often necessary in cases of gestational diabetes (Zolghadri et al. 2008). Diabetic pregnant women whose glucose levels are well controlled are not at significantly higher risk of miscarriage than pregnant women without diabetes mellitus. On the other hand, the likelihood of miscarriage and malformations of the child is clearly increased in pregnant diabetics with poor metabolic control during the first trimester, i.e. with high levels of glucose and glycosylated hemoglobin (Deutsche Diabetes-­Gesellschaft 2014). Hypo- or hyperthyroidism are often mentioned as possible causes of sporadic or recurrent pregnancy losses, but there are also reports to the contrary in the literature. Although a thyroid dysfunction is to be expected in only up to 2% of women with recurrent pregnancy loss, an examination of the thyroid function in these patients seems to be justified with regard to the easy correctability of these dysfunctions and also because of a possible worsening during pregnancy. The lower normal range should be considered as the target TSH level, even

10

though only small improvements in miscarriage rates have been shown in the literature (Reid et  al. 2010). An association of pregnancy loss with the presence of thyroperoxidase (TPO) antibodies has been shown, but it is unclear whether therapeutic measures other than correction of thyroid function are required (Prummel and Wiersinga 2004). A reduction of the postpartum thyroiditis rate could be achieved by the additional use of selenium (Reid et al. 2010).

Psychosocial Factors The psychological trauma as a consequence of one or even several pregnancy losses is widely underestimated. The awareness of having lost a pregnancy is intensified by the possibilities of early sonographic diagnosis. After experiencing multiple pregnancy losses, the fear of recurrent pregnancy losses is all too understandable, which is why a high incidence of reactive depression and anxiety is found in couples with recurrent pregnancy loss. Although psychological factors are unlikely to be the cause of repeated pregnancy loss, neglecting these aspects is potentially detrimental. Several studies have demonstrated a high rate (75%) of successful pregnancies in patients with recurrent pregnancy loss solely through “tender loving care” in conjunction with short-term clinical and sonographic monitoring (Stray-­ Pedersen and StrayPedersen 1984; Rai et al. 1996), although the high spontaneous s­ uccess rate must be taken into account when evaluating these measures.

10.2 

Recurrent Pregnancy Loss

Ruben KuonKilian Vomstein, and Bettina Toth

10.2.1 

Introduction

While about 30% of all women experience a spontaneous pregnancy loss in their lifetime, the incidence of recurrent (habitual) sponta-

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neous pregnancy loss (RPL) is 1–3% depending on the definition used. The WHO defines the presence of RPL after three consecutive miscarriages before the 20th week of gestation, the American Society for Reproductive Medicine already speaks of RPL after two consecutive miscarriages. Established risk factors include endocrine, anatomical, infectiological, genetic, psychological, hemostaseological and immunological factors (. Fig. 10.7). After standardized diagnostics, an explanatory cause can be identified in about 50% of affected women; the other half of RPL remains unclear, which is why the establishment of new diagnostic and therapeutic approaches is urgently needed. In the following section, we will first discuss the individual established risk factors. We then focus on potential new risk factors such as chronic endometritis (CE) and the presence of peripheral and uterine natural killer cells (pNK and uNK cells) in patients with RPL.  

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10.2.2 Established Risk Factors

Endocrine Dysfunctions Endocrine causes of RPL include luteal phase insufficiency, thyroid dysfunction, metabolic syndrome including obesity, and PCO syndrome (PCOS) (. Fig. 10.8). The diagnosis of luteal phase insufficiency includes determining the length of the menstrual cycle, measuring the progesterone level in the luteal phase and, if necessary, performing an endometrial biopsy to detect secretory transformation of the endometrium. To date, there are no studies that have demonstrated a clear association between the occurrence of RPL and the diagnosis of luteal phase insufficiency. Thyroid dysfunctions include both manifest hyperthyroidism and manifest hypothyroidism, both of which are associated with the occurrence of miscarriages (Anselmo et al. 2004). At present, however, the data are unclear as to what extent latent  

23.10%

46.60% Hemostaseology Autoimmunology Genetics Anatomy 7.80%

46.60%

Endocrine factors

5.80%

..      Fig. 10.7  Incidence of individual established risk factors in RPL patients (own previously unpublished data). The respective risk factors include: Endocrine factors: thyroid disease (hypo−/hyperthyroidism, antiTPO antibodies) and luteal phase insufficiency; Autoimmunology: antinuclear antibodies (ANA titer >1:160), anti-cardiolipin antibodies (ACL IgG/IgM),

anti-ß2-glycoprotein IgG/IgM, lupus anticoagulant; Hemostaseology: Factor V Leiden, prothrombin or methyltetrahydrofolate reductase (MTHFR) mutation, protein C/S deficiency, antithrombin deficiency; Genetics: parenteral chromosomal disorders; Anatomy: uterine septum

231 Disorders of Early Pregnancy and Pregnancy Loss

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..      Fig. 10.8  PCO-typical ovaries in transvaginal ultrasound. The classic pearl string-like arrangement of the follicles is depicted. (With kind permission of Prof. Dr. Brezinka, MUI Innsbruck)

hypothyroidism (elevation of TSH concentrations in the presence of normal thyroid hormone concentrations) is also associated with (recurrent) miscarriages. In principle, the Endocrine Society recommends an upper TSH value of 2.5 mU/l (Abalovich et  al. 2007), which should already be reached pre-­conceptually. The pathophysiological significance of the metabolic syndrome including obesity and PCOS for the occurrence of RPL is often correlated as there is overlap. Even though the data from international studies are inconsistent, the BMI should be determined in patients with RPL and a metabolic syndrome should be investigated for, if the BMI is ≥30 kg/m2. Prior to a new pregnancy, weight reduction should already be attempted.

ment (Raga et al. 2009). Therefore, surgical hysteroscopy with septum resection is recommended prior to the onset of a new pregnancy. Similarly, there is evidence that a so-called uterus unicornis is associated with the occurrence of RPL or late pregnancy losses or preterm births (Ozgur et al. 2017), however, therapeutic approaches are lacking. The diagnosis of congenital or acquired uterine malformations is performed using (3D) vaginal sonography or diagnostic hysteroscopy. Acquired anatomical disorders include intrauterine adhesions, which may occur mainly after dilation and curettage or infection, as well as myoma and polyps. Intrauterine adhesions can be detected by diagnostic hysteroscopy and should be removed. Depending on their location, myoma/ Anatomical Malformations fibroids are classified as submucous, intraAnatomical causes include both congenital mural and subserous (see also FIGO classifi(such as uterine malformation or other uro- cation). Submucosal and large intramural genital malformations) and acquired (such myoma, which compress the uterine cavity, as intrauterine adhesions, polyps, myoma) are associated with the occurrence of malformations. RPL. However, data is sparse and inconsisThe incidence of uterine malformations tent. Nevertheless, in the presence of subin patients with RPL varies between 3–25% mucous or intramural myoma compressing in studies (Salim et  al. 2003; Sugiura-­ the uterine cavity, a pre-conceptional hysOgasawara et  al. 2011), with uterus septus teroscopic or laparoscopic enucleation can being the most common. be performed. Pathophysiologically, it is assumed that The same applies to intracavitary polyps. the septum leads to a disturbance of vascu- As in cases of myoma, there is no internalarization in the course of placental develop- tional consensus on the size or number of

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polyps that are relevant for the occurrence of RPL and therefore have to be removed preconceptionally. Nevertheless, the removal of polyps with histological examination is recommended from a size of 15  mm to exclude a (rare) malignant degeneration.

Infections

10

Especially in the presence of recurrent late pregnancy losses with premature rupture of the membranes, premature labor and opening of the cervix as well as amniotic infection syndrome, vaginal infections should be excluded in the patient. In the course of a gynecological examination, both a smear from the vagina and a bacteriological s­ ample should be taken. In the event of an infection, antibiotic treatment should be administered as proven in the antibiogram. For prophylactic purposes, vaginal suppositories containing Döderlein bacteria can be used before pregnancy.

Chromosomal Disorders After the sperm has fertilized the oocyte, a number of developmental steps are necessary for proper embryonic development to occur. These developmental steps are highly error-prone, which leads to frequent (unnoticed) miscarriages or failure of embryo implantion in a woman’s lifetime (Laurino et al. 2005). Furthermore, the probability of embryonic or fetal chromosomal maldistribution shows a strong correlation with maternal age, and hereditary genetic disorders may also be present. Concerning chromosomal aberrations in miscarriage tissue, trisomy 16 is identified most prevalently, however, trisomy 22, triploidy and monosomy X (Turner syndrome) are also found frequently. Only about 4–5% of RPL couples are diagnosed with  a (balanced) chromosomal change in at least one of the partners. Not all  international guidelines recommend a standardized chromosome analysis in the affected couples (De Braekeleer and Dao

1990). The guidelines of the German, Austrian and Swiss Society of Gynecology and Obstetrics (DGGG, OEGGG, SGGG) for the diagnosis and treatment of RPL currently recommend a human karyotyping of the affected couple or the product of conception. In principle, prior to any genetic diagnosis, an information on the planned genetic diagnostics should be performed by a qualified physician, including the written consent of the couple or patient concerned, in accordance with the genetic diagnostics law of the respective country. If one of the partners of the affected couple is diagnosed with a balanced chromosomal abnormality, the risk of a pregnancy loss as well as the risk of giving birth to a child with a chromosomal abnormality depends on the chromosomes affected. There is no causal therapy for the treatment of maternal or paternal chromosomal abberations or for the prevention of unbalanced chromosomal rearrangements. However, pre-implantation genetic diagnosis (PGD) can be used to perform a genetic diagnosis of the embryo before transfer. In PGD, a distinction is made between the examination of already known genetic diseases (e.g. monogenic disease or balanced chromosomal aberration) and the screening for embryonic chromosomal aberrations. Internationally, the terms “preimplantation genetic testing” (PGT) and “preimplantation genetic diagnosis” (PGD) are increasingly used. Sole screening, on the other hand, is referred to as “preimplantation genetic screening” (PGS). In principle, PGD is performed on trophectoderm cells; the examination of blastomeres has lost priority. The prerequisite for performing PGD in Germany is regulated in the Embryo Protection Act and is only permitted at PGD centers approved for this purpose. In addition, a positive vote by an ethics committee must be obtained before any PGD is performed in Germany.

233 Disorders of Early Pregnancy and Pregnancy Loss

In contrast, no ethical vote is required in advance for polar body diagnostics (PBD). However, PBD only permits an inference to maternal chromosomal maldistributions, the paternal side remains unclear. In the case of PGD (including PBD), the affected patient undergoes hormonal stimulation as part of assisted reproductive therapy (ART), which is costly and time-consuming and may involve risks of side effects. A PGS enables the selection of genetically healthy embryos. However, contrary to expectations, no improvement in the live birth rate in patients with RPL and PGS has been observed in international studies to date, so that the DGGG/OEGGG/SGGG guideline does not currently recommend the performance of a PGS in RPL patients.

10

already physiologically present increased procoagulatory systems in pregnancy. As a consequence, microthrombi could form in the placental bed, which in turn cause a reduced blood flow to the placenta and thus reduce supply of the embryo or fetus. Up to 15% of the Caucasian population exhibit one of the thrombophilic parameters mentioned (Roberts et al. 2009), so that it is evolutionarily questionable why such thrombophilias have been passed on over such a long period of time. One possible explanation could be the positive effect of a procoagulant tendency: It is possible that the patient suffers less blood loss peripartum, which may have been an evolutionary advantage until just a few years ago. Due to the inconsistent data situation, international guidelines do not recommend Psychological Factors screening for maternal hereditary thromboExperiencing RPL is a perturbing experi- philia outside of studies. However, thromence for the couple affected and can lead to bophilia screening should be performed if severe traumatization. Particularly in the risk factors are present (family history of case of pre-existing psychiatric illnesses, thrombosis, own history of thrombosis). close supervision together with a psycholoThe administration of low-molecular-­ gist or psychiatrist should be initiated pre-­ weight heparins (LMWH) in patients with conceptually. RPL and the presence of a hereditary thrombophilia is recommended in international Hemostasiological Factors guidelines, but only in risk constellations and The clarification of hereditary thrombophil- primarily for the reason of maternal thromias in patients with RPL is currently the sub- bosis prophylaxis, but not for the prevention ject of international controversy. of a new pregnancy loss. This recommendaClassical hereditary thrombophilias, tion is mainly based on the results of numerwhich have been investigated in numerous ous international studies, which could not international studies in RPL patients, prove a benefit with regard to the live birth include mutations in the factor V Leiden rate after the administration of LMWH, nei(FVL; c.1601G > A in F5, rs6025) or pro- ther in the presence of thrombophilia nor in thrombin gene (G20210A) (PT; c.*97G > A so-called idiopathic RPL (without identificain F2, rs1799963) as well as polymorphisms tion of an established risk factor). in the methylenetetrafolate reductase gene In special risk constellations such as the (MTHFR C677T or c.665C > T). presence of an antithrombin deficiency, a Furthermore, a deficiency of antithrombin, homozygous FVL mutation or a combined protein C, protein S, protein Z or factor XII heterozygous FVL and PT mutation, howas well as an increased concentration of fac- ever, interdisciplinary care should be protor VIII or lipoprotein (a) (Toth et al. 2008) vided together with hemostaseologists, are associated with RPL. Pathophysiological obstetricians and, if necessary, neonatoloconsiderations assume that hereditary gists in addition to (therapeutic) heparinisathrombophilia has an additive effect on the tion of the pregnant woman.

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Immunological Factors Immunological risk factors for RPL include a variety of allo- and autoimmunological factors. However, only the antiphospholipid syndrome (APS) is established as an immunological risk factor for RPL in international guidelines. By definition (see following overview), laboratory criteria must be fulfilled in addition to clinical criteria for the diagnosis of APS. APS is present in about 2–15% of patients with RPL (Branch et al. 2010). In order to diagnose an APS, laboratory testing of antiphospholipid antibodies in medium to high range (>99th percentile measured in normal subjects) has to be redone 12 weeks after the initial positive blood test (Miyakis et al. 2006a, b). Diagnostic Criteria for Antiphospholipid Syndrome (Miyakis et al. 2006a, b)

10

Clinical Criteria: 55 ≥1 venous or arterial thrombosis 55 One or two unexplained pregnancy losses in morphologically normal fetuses >10th week of gestation 55 ≥3 pregnancy losses 160/110 mmHg and/or the proteinuria values of >3 g/dl protein, this is referred to as severe preeclampsia. Definition of Preeclampsia A revised definition of pre-eclampsia came from the ISSHP in 2014 and includes the following criteria: 55 De novo hypertension after 20 weeks gestation together with one or more of the following additional medical conditions: –– Proteinuria –– Urine protein/creatinine >30 mg/mmol (0.3 mg/mg) or –– >300 mg/day or –– at least 1 g/L [“2+”] in a urine dipstick measurement –– Other maternal organ dysfunctions –– Renal insufficiency (creatinine >90 μmol/l; 1.02 mg/dl) –– Liver involvement (elevated transaminases—at least two times the upper limit of normal and/or pain in the upper right quadrant of the abdomen or epigastric abdominal pain) –– Neurological complications (examples include eclampsia, altered mental status, blindness, stroke) –– Hematologic complications (thrombocytopenia—platelet counts >So far, it is still open whether the link between preeclampsia and increased later risk of obesity, diabetes and cardiovascular disease and death is direct or indirect. There could be a direct link, and preeclampsia during pregnancy increases later risk. However, it could also be that the woman has an undiagnosed pre-existing condition which then leads to preeclampsia during the “stress test pregnancy.” Thus, even without pregnancy and thus without preeclampsia, this woman would have an increased risk later in life.

The long-term effects not only affect the mother, but also the child is epigenetically reprogrammed by stress during pregnancy (Choudhury and Friedman 2012). This direct fetal programming is associated with an increased risk of children developing obesity and diabetes in adolescence (von Ehr and von Versen-Höynck 2016; Godfrey and Barker 2001).

Pathophysiology Although massive efforts have been made in recent decades to elucidate the etiology of preeclampsia, this syndrome still remains

what it has been for decades: a pathology of hypotheses. A few points are clear and unambiguous: it takes the placenta, but not the fetus for preeclampsia. Molar pregnancies without a fetus can still develop preeclampsia. Once the placenta is born, the woman’s symptoms disappear. Due to the many risks that can influence the development of preeclampsia, it is now assumed that it is a multifactorial event in which both the placenta and the predisposition of the woman play decisive roles. The interplay of these two factors determines the development of preeclampsia as well as its severity: early or late onset, mild or severe, etc.

 isk Factors for the Development R of Preeclampsia A variety of risk factors have now been described that increase a woman’s risk of developing preeclampsia during pregnancy. These include in particular: 55 previous preeclampsia, especially if it was severe or early onset preeclampsia (3 pregnancies the risk increases beyond that of the first pregnancy, 55 First paternity (primipaternity or alternate paternity or a period of >5  years between two pregnancies with the same father),

247 Placental Insufficiency/Placenta-Associated Diseases

55 Short period of sexual intercourse (85%. At the same time, there are clear differences between early and late onset preeclampsia. Late onset preeclampsia can generally be characterized as follows: 55 The child displays normal height and weight. 55 Blood flow in the uterine arteries is normal or only very slightly altered. Thus, there are no significant changes in Doppler ultrasound or pulsatility index. 55 Blood flow in the umbilical arteries is unchanged. 55 Women with increased placental mass/ surface area (diabetes mellitus, multiple pregnancies, anemia, high altitude) are at increased risk for developing late onset preeclampsia. Most early onset preeclampsia cases, on the other hand, have the following characteristics: 55 The child is often too small and has a growth restriction. 55 Blood flow in the uterine arteries is often altered. With this there are clear changes in Doppler ultrasound and increased pulsatility index. 55 Increased peripheral resistance in the placental vessels could be a reason for the change in blood flow in the umbilical arteries. This may involve changes in blood flow in these vessels with flow still present (PEDF, preserved end-diastolic

11

flow). Blood flow may stop at the end of diastole (AEDF, absent end-diastolic flow) or blood may even flow backwards through the umbilical arteries at the end of diastole (REDF, reversed end-­ diastolic flow). 55 Trophoblast invasion is insufficient, especially with regard to endoarterial invasion into the spiral arteries. However, it must be made clear that the characteristics “specific” to early onset preeclampsia are not specific to this syndrome! All the characteristics listed above for early onset preeclampsia also apply to early fetal growth restriction (FGR), which is not associated with maternal symptoms. The typical characteristics of early FGR are: 55 Of course, a child with a growth restriction, 55 Inadequate trophoblast invasion, especially into the spiral arteries, 55 Change in blood flow through the uterine arteries (increased pulsatility index), 55 Changes in blood flow through the umbilical arteries (PEDF via AEDF to REDF). Since the two early syndromes, early onset preeclampsia and early onset FGR often occur in parallel, the question must be asked whether they are directly associated with each other or whether, under certain circumstances, they occur either alone or in parallel. There is also an attempt in the literature to explain that one syndrome (early onset preeclampsia) is responsible for the second syndrome (early FGR). Of course, this can only apply to cases where both occur in parallel. Unfortunately, clarification is not yet in sight.

Presentation and Falsification of the Outdated Hypothesis on the Etiology of Preeclampsia Interestingly, a hypothesis on the etiology of preeclampsia, which uses the deficient invasion of the extravillous trophoblast as a

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basis, has been described in the scientific literature for decades. It has long since been disproved by a large number of studies. The continued citation as one of the most important hypotheses demonstrates the importance of a critical examination of hypotheses. At the same time, it also shows that the falsification and rejection of a hypothesis as the most important tool of science does not necessarily lead to its abandonment. However, since the said hypothesis is still being circulated today, here is a brief summary of the chronological sequence of events described in this hypothesis: 55 First trimester: a previously undescribed deleterious effect on the extravillous trophoblast. 55 First trimester: Deficient invasion of the extravillous trophoblast with a markedly reduced invasion of the spiral arteries in the first and second trimester. 55 Second trimester: reduced flow of maternal blood into the intervillous space of the placenta. 55 Second and third trimesters: placental hypoxia or events of hypoxia followed by reoxygenation of the placenta. 55 Second and third trimester: hypoxic damage to the villous trophoblast. 55 (Second and) third trimester: release of placental factors of the syncytiotrophoblast (such as sFlt-1 and PlGF) into the maternal blood. 55 (Second and) third trimester: maternal inflammatory reaction due to these placental factors and development of the mother’s clinical symptoms. This was a conclusive hypothesis over 10  years ago that could explain many aspects of preeclampsia. It has since been refuted in many places. It is therefore all the more surprising that it is still cited and supported. In the following, two events crucial to this hypothesis will be considered in more detail.

Deficient Trophoblast Invasion and Preeclampsia In the scientific literature in connection with the pathophysiology of preeclampsia in general, one almost always finds the change in the extravillous trophoblast that is only known for early onset preeclampsia: deficient trophoblast invasion. Considering that early onset preeclampsia accounts for only 10–15% of all preeclampsia cases, it must be asked how this came about and how the etiology of the other 85–90% of preeclampsia cases is to be explained. Since early onset preeclampsia has proportionally the highest number of cases with severe complications and is also clinically extremely relevant due to the early gestational age at birth, such early onset preeclampsia cases have received and continue to receive significantly more attention and thus more research than the late cases—even though the latter occur five to six times more frequently. Early onset preeclampsia cases are often associated with FGR, which is why studies of early onset preeclampsia have led to the erroneous conclusion that these changes apply to all preeclampsia cases. However, since pure FGR cases also show the symptoms described above without the mother becoming symptomatic, it must be critically questioned what the true pathophysiology of early onset preeclampsia is. This criticism is supported by studies on the prediction of preeclampsia using uterine Doppler ultrasound as a surrogate for deficient trophoblast invasion. Measurements of an elevated pulsatility index in the uterine arteries at 11 + 0 to 13 + 6 weeks gestation predicted preeclampsia (all cases) in only 40% with a 10% false positive rate (Nicolaides et al. 2006). Another study also showed a clear picture: here, too, blood flow in the uterine arteries was measured with Doppler ultrasound at the 11th–14th week of pregnancy. Based on the corresponding data, the authors were able to predict 21%

249 Placental Insufficiency/Placenta-Associated Diseases

of all preeclampsia cases and only 33% of early onset preeclampsia cases. This could be expected against the background described above. At the same time, however, the authors were able to predict 100% of all early onset FGR cases (Pilalis et al. 2007). Such and further studies increase the doubt that the deficient invasion of the extravillous trophoblast is causally involved in the etiology of preeclampsia. The dysregulation of the extravillous trophoblast seems to result mainly in fetal growth restriction, which occurs even in the absence of preeclampsia. Thus, the above-mentioned ­ characteristics of early onset preeclampsia would be due to early onset FGR. And thus, up to this point, there would be no specific characteristics of early onset preeclampsia that could distinguish it from late onset preeclampsia—except for its co-occurrence with early onset FGR, which usually necessitates early delivery.

 eficient Placental Perfusion, D Placental Hypoxia and Preeclampsia As already described in 7 Chap. 1, during the first trimester there is only a maternal plasma flow passing through the placenta. Thus, only oxygen physically dissolved in the fluid with a partial pressure of 80% of all preeclampsia cases. Here, the hypothesis described above does not apply at all.

251 Placental Insufficiency/Placenta-Associated Diseases

Possible Explanation of the Etiology of Preeclampsia As described above, the placenta is inevitable for the development of preeclampsia. At the same time, the placenta cannot be considered alone here, but must always be analyzed in the context of its environment. Only the interaction between placenta, release of placental factors into the maternal circulation, and maternal response to these factors will determine whether or not a pregnant woman will develop preeclampsia. Therefore, the effect of each side must be investigated in order for symptoms to become clinically relevant (Huppertz 2008). Given the close interactions between maternal and fetoplacental factors during pregnancy, at least three scenarios can be considered. These different scenarios can explain the different types and manifestations of preeclampsia, the different times of onset and the different effects on mother and child in later life (. Fig. 11.1):  

z Scenario 1a: A Healthy Mother with a Malfunction of the Placenta in the Villous Trophoblast

During normal pregnancy, the syncytiotrophoblast releases factors by secreting substances in a controlled manner or by releasing them as syncytial knots via apoptotic processes (Huppertz et al. 2006; Huppertz 2008, 2010). In this scenario, dysfunction of the syncytiotrophoblast results in subcellular material entering the maternal circulation through necrosis and aponecrosis (Huppertz et al. 2006; Huppertz 2008, 2010). This release of subcellular particles such as micro- and nanoparticles (Johansen et al. 1999; Redman and Sargent 2000) systemically activates and permanently damages the maternal endothelium (Goswami et al. 2006). Dysfunction of the placenta and specifically the syncytiotrophoblast leads to the ongoing release of factors that can activate and/or damage the maternal vasculature and thus induce preeclampsia.

11

The extent of this damage can be directly correlated with the surface area of the placenta: The greater the placental surface area (e.g., in large placentas [diabetes], in multiple pregnancies, or pregnancies at high altitude), the greater the amount of non-apoptotic factors released. In addition, the maternal defense system is overloaded above a certain amount of factors, and thus damage to the maternal endothelium is possible even faster. If the maternal defense system can withstand the amount of placental factors over a longer period of time, the clinical symptoms of preeclampsia will develop late (late onset preeclampsia). Since this implies that the maternal endothelium also suffers damage late in pregnancy, predictive markers that represent damage to the endothelium (such as the angiogenic factors sFlt-1 and PlGF) will not show changes before the onset of clinical symptoms. Moreover, angiogenic factors are more related to FGR (Nicolaides et  al. 2006; Pilalis et  al. 2007). At the same time, placenta-specific predictive markers such as PP13 show changes in their release pattern from the syncytiotrophoblast already in the first trimester (Huppertz et al. 2008). In this scenario, a normal-weight child usually develops without growth restriction. However, damage to the syncytiotrophoblast can also negatively affect the absorption of nutrients, so that it cannot be ruled out that at least late effects are possible for the child in adulthood. After birth, the mother should recover completely and not suffer any long-term consequences. z Scenario 1b: A Healthy Mother with a Malfunction of the Placenta in the Extravillous Trophoblast

If there is a defect of the extravillous trophoblast in the placenta, the release of factors from the syncytiotrophoblast is not altered. In this case, however, there is

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Predisposed mother

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Normal risk for mother, but Normal risk for mother and child later in life increased risk for child later in life

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Reduced growth of the fetus (FGR)

Increased risk for mother, but normal risk for the child later in life

Increased risk for mother and child later in life

..      Fig. 11.1  Schematic representation of the scenarios that can lead to preeclampsia and/or fetal growth restriction (FGR). Scenario 1 describes the effects on mother and child in the case of a healthy mother and a malfunction in the placenta. Scenario 2 shows the

effects on mother and child in a pre-­diseased mother and a healthy placenta, while Scenario 3 shows the effects on mother and child in a pre-diseased mother and a malfunctioning placenta

deficient invasion of the extravillous trophoblast with subsequent altered delivery of maternal blood to the placenta. One effect of the increased flow velocity of the m ­ aternal

blood is an increase in peripheral resistance in the small vessels of the placental villi. This increased resistance can lead to changes in fetal blood flow to the placenta, resulting

253 Placental Insufficiency/Placenta-Associated Diseases

in poorer delivery of both nutrients and oxygen to the fetus. Evidence of arrest or reflux of blood flow in the umbilical arteries at the end of diastole (AEDF or REDF) reveals corresponding serious consequences for the fetus. In these situations, a hypoxic fetus may be associated with a hyperoxic placenta: The same amount of blood (and therefore oxygen) flows into the placenta from the maternal side. Due to the increased flow velocity in the placenta and the damaged villous surface, the placenta absorbs less oxygen and passes less oxygen on to the fetus. Thus, the fetus develops a hypoxia while there is a higher partial pressure of oxygen in the placenta compared to the normal placenta. Hyperoxia occurs in the placenta in these cases of FGR. The following scenario has not yet been clarified and can only be depicted hypothetically: The alteration of the maternal blood flow in the placenta and the damage to the villous structure may subsequently also lead to damage of the villous surface and thus to a release of subcellular material from the syncytiotrophoblast. Thus, in this scenario, the deficient trophoblast invasion could cause FGR of the fetus and subsequently also induce preeclampsia. In this scenario, a child develops with growth restriction and it cannot be ruled out that late effects in adulthood are possible. If the mother does not develop preeclampsia, she should not suffer any long-term consequences. z Scenario 2: A Predisposed Mother with a Normal Placenta

In this scenario, the placenta releases the normal portfolio of fragments, factors and molecules into the maternal circulation. However, pre-existing damage in the mother implies that the “pregnancy stress test” is not without complications for the mother. The pre-existing damage may be to the

11

woman’s defense system, to the endothelium itself, or to signaling cascades that regulate renal function and blood pressure (e.g., chronic hypertension, antiphospholipid syndrome, etc.). Ultimately, this damage causes the overall system of a pregnant woman to respond inadequately to placental factors, resulting in the development of clinical symptoms of preeclampsia. This is true even if the quality and quantity of placental factors are within the normal range. Also in this scenario, the extent of preeclampsia can be correlated with the surface area of the placenta. The greater the placental surface area, the greater the amount of apoptotic factors released. Depending on the damage to the maternal system, the system may become overloaded early (early onset preeclampsia) or only towards the end of pregnancy (late onset preeclampsia), leading to the expression of clinical symptoms. For predictive markers released from the maternal side, changes could occur early and thus indicate an increased risk for the woman. At the same time, no difference will be seen in placenta-specific markers, as the placenta does not show any changes. In this scenario, most often a normal weight child develops without growth restriction and without expected late effects in adulthood. This looks different in the mother. Since the woman already suffers from a subliminal and subclinical disorder, this can lead to a higher risk of cardiovascular disease later in life. z Scenario 3: A Predisposed Mother with Placental Dysfunction

This scenario contains the most severe cases, as the combination of defects on both sides can lead to severe symptoms in the child, but especially in the mother. On the placental side, different sub-­ scenarios can be described. 55 If the villous trophoblast alone is affected, damage to the villous surface

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will occur, resulting in the release of subcellular particles from the syncytiotrophoblast into the maternal blood. These necrotic particles will then encounter an already pre-damaged vascular system of the mother. Thus, the clinical picture of preeclampsia will develop early in these women without the need for growth restriction of the fetus. Such cases should be predictable with placenta-specific markers such as PP13 as well as with vascular-associated markers such as sFlt-1 and PlGF. 55 If not only the villous but also the extravillous trophoblast is affected in the placenta, deficient invasion also occurs (Huppertz 2011). In these cases, the syncytiotrophoblast releases necrotic factors and the deficient invasion alters blood flow through the placenta (Burton et al. 2009). This can lead to growth restriction of the child, but in addition can also increase the extent of syncytial changes. In this worst case scenario, early severe preeclampsia of the mother with associated early onset FGR of the child may occur. These cases can be detected with all predictive markers, both placenta-specific (Chafetz et al. 2007) and angiogenic markers (Schaarschmidt et al. 2013). In this third scenario, mothers are at significantly increased risk of cardiovascular disease and other conditions later in life. In cases with FGR, the children will also suffer long-term damage from fetal programming (Longtine and Nelson 2011; Hogg et  al. 2013). All these different scenarios make it clear that the origin and etiology of preeclampsia will not be clarified for a long time yet. New approaches and explanatory models must be found through appropriate research actions in order to finally come closer to clarifying the origin of preeclampsia.

11.2 

 GR: Diagnostics and F Management

Ulrich Pecks

Fetal growth restriction (FGR) occurs when a fetus is unable to reach its genetically predetermined growth potential. The reduced growth rate results in a low birth weight usually below the tenth percentile. Antenatal sonography and risk history help in correct diagnosis in differentiation from constitutionally small-for-gestational-­ age (SGA) fetuses and in further assessment of outcome. The challenge for the obstetrician is to determine the optimal time of delivery in order to avoid fetal death while minimizing morbidity and mortality associated with preterm birth. 11.2.1 

Terminology and Definition

The terminology used in the literature for children born small or light is often not very clear-cut. “Low birth weight” (LBW), “small for gestational age” (SGA) and “intrauterine or fetal growth restriction” (FGR/FGR) are often used synonymously. >>A birth weight >Fetal growth restriction (FGR) occurs when a fetus is unable to reach its genetically predetermined growth potential during pregnancy.

Strictly speaking, the term retardation (lat. “retardare,” to delay), which is often found in the literature, is not correct here, since in the narrower sense there is no delay but a restriction (lat. “restringere,” to restrict) of fetal growth.

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Fundus stand in cm

42 40

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GROW Chart © Gestation Network

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24 25 26 SSW

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..      Fig. 11.2  Fetal weight percentile curves of two patients of different origin, body measurements and parity at 36 + 4 weeks gestation with female fetuses and an estimated fetal weight of 2500  g. Ms. Short (bottom) shows adequate fetal growth in the 40th percentile,

35 36 37 38 39 40 ET

41 42

500 0

while Ms. Tall (top) has an SGA fetus in the fifth percentile. (Personalized percentile curves courtesy of Prof. Jason Gardosi, Perinatal Institute Birmingham, UK; © Gestation Network—7 www.­gestation.­net)  

257 Placental Insufficiency/Placenta-Associated Diseases

11.2.2 

Epidemiology

In Germany, approximately 53,000 children (7.22%) were born with LBW in 2015. By definition, 10% of all children are born as SGA (> A pathological Doppler of the umbilical arteries is the best discriminating prognostic factor with regard to fetal outcome.

100

% of FGR diagnoses

(PIV) increases to the point of flow reversal during the so-called a-wave, in which atrioventricular inflow is reflected (Kiserud et al. 2006). Once the fetus’ adaptive mechanisms to its nutritive insufficiency are exhausted and the fetus is no longer able to maintain this central redistribution, signs of the so-called “late cardiovascular response” occur, with a decrease in cardiac ejection, multiphasic venous patterns, and possibly a normalization of the middle cerebral artery (Ferrazzi et al. 2002). With increasing hypoxia and deterioration of the acid-base balance, the fetus responds with behavioral mechanisms. There is a decrease in overall activity and respiratory movements to complete inactivity with abolished flexion of the extremities (Manning 2002; Frøen et  al. 2008). The fetal cardiac tone curve shows nonreactivity, which can be seen on cardiotocography (CTG) and Oxford CTG (Arabin et  al. 1988; Visser et al. 2016). On the basis of different clinical courses and compensation patterns depending on the gestational age, a distinction is increasingly being made between an early (“early onset”) and a late (“late onset”) form of FGR. Early forms of FGR are more often associated with an increased resistance index (RI) of the umbilical arteries up to end-diastolic absent or reversed flow (ARED flow) (Baschat 2011). Using a cohort of 656 FGR singletons, Savcheva et  al. found that the 32nd week of gestation best distinguished between an early and late form of FGR using Doppler sonographic parameters of the umbilical arteries (Savchev et al. 2014). This is consistent with observations from our own cohort of 95 FGR singletons (. Fig. 11.4).

11

50

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25-28

29-32 33-36 week of gestation

37-41

..      Fig. 11.4  Classification of FGR cases with and without ARED flow of the umbilical arteries. Collective of 95 cases who participated in a study at the University Women’s Hospital of RWTH Aachen in 2006–2014 (Pecks et al. 2012, 2016). Shown is the percentage of patients with ARED (black bars) and without ARED (white bars) in their gestational age group

11.2.5 

Outcome of the Child

It is undisputed that FGR can be held responsible for the majority of intrauterine fetal deaths (IUFD) (Hübner et  al. 2015). According to an English study of 2625 stillbirths, FGR accounted for 42% of IUFD.  In comparison, congenital anomalies accounted for the second largest proportion of all IUFD at 17% (Gardosi et al. 2014). Based on a survey of infants born in the United States in 2005, the risk of IUFD in SGA infants increases as the percentile group decreases. Using population-based growth curves, fetuses 10th percentile reference group (Pilliod et  al. 2012). At a fetal weight 90% can be achieved. A similar method for the isolation of decidual macrophages in the third trimester was also described by Narahara (Narahara et al. 1993) and has been continuously developed since then (Narahara et  al. 2003). In this method, the first digestion with protease XIV and dispase is bypassed; instead, an additional enrichment of mononuclear cells in a Ficoll-Hypaque gradient was introduced before the Percoll density gradient. Although viability of >90% is also achieved here and the yield of about 15 × 106 cells/g tissue is remarkably high, this method isolates a rather low amount of pure macrophages, as illustrated by the proportion of CD14-positive cells (90% and purity of >95% can be achieved with such treatments (Wilson et al. 1983). Second, macrophages, in their capacity as immune cells, have numerous surface receptors that can be exploited in purification. One technique for this is “rosetting,” in which antibodies to HBC proteins are anchored in the membrane of erythrocytes, which are then incubated with an HBC cell suspension. HBCs bind to the red cells, while other cells remain unbound and can be separated by centrifugation. The erythrocytes then still need to be lysed to obtain pure HBCs (Sutton et  al. 1989). Another common option is purification by magnetic beads to which an antibody to the macrophage marker CD68 has been coupled (Cervar et  al. 1999). Both rosetting and purification via CD68 involve “positive immunoselection,” i.e. HBCs are bound via an antigen and all other unbound cells are removed by washing. Since positive immunoselection may result in undesirable proinflammatory activation of macrophages, which could affect results especially in functional assays, recent protocols for isolation of HBCs use negative immunoselection, i.e. all non-HBC cells are bound via antibodies on magnetic beads and removed by a magnet, while HBCs remain in suspension. For negative selection, e.g. antibodies against Epidermal Growth Factor Receptor (EGFR) are used

on cytotrophoblasts, which are then removed (Wetzka et  al. 1997), leaving HBC, which then have a purity of >90% (measured on CD68+ cells). In a second step, fibroblasts can also be removed using an antibody against CD10 (Tang et al. 2011). With this method, the purity increases to almost 99% (measured on CD163+ cells) with a yield of approx. 1.5 × 106 cells/g tissue. There is also the possibility of isolating Hofbauer cells from first trimester placental tissue, the protocol for this is very similar to the methods used for isolation from term tissue: digestion is unnecessary, the tissue is very finely minced and applied sequentially to Ficoll Hypaque and Percoll gradients, then the cells are enriched according to their surface molecules by rosetting (Zaccheo et al. 1989). However, the starting material is the limiting factor; 10–15 tissue samples must be collected and pooled to perform an experiment (Zaccheo et  al. 1989). Immediately after isolation, Hofbauer cells look mostly round and, characteristic for these cells, show a large number of vacuoles (. Fig.  15.3a). After about 2  days in culture, the cells begin to elongate and form projections (. Fig. 15.3b). They are capable of phagocytosis and, for example, of taking up cholesterol (. Fig. 15.3c). In addition to the relatively laborious isolation, a disadvantage is that the cells can only be kept in culture for 7–10 days, and there is no possibility of freezing the cells and using them at a later time point. In addition, HBCs are non-proliferative, so the yield at the end of isolation is a limiting factor in the design of further experiments. However, as human primary cells, HBCs have high plasticity and thus can be used in a variety of assays to investigate their role in placental angiogenesis (Loegl et al. 2016), vertical transmission of infectious diseases (Johnson and Chakraborty 2012; Simoni et  al. 2017), or inflammatory diseases of the placenta (Ben Amara et al. 2013; Kim et al. 2008).  

 

 

343 Research Aspects and In Vitro Models

a

15

b

c

..      Fig. 15.3  a–c Hofbauer cells (HBCs) in culture. a 24 h after isolation HBCs are still small and round. b 96 h after isolation HBCs are already elongated and

15.5 

Placenta Ex Vivo Perfusion

The human placenta is delivered immediately after delivery of the child and is thus promptly available for ex vivo examinations of the tissue. Ideally, the structural integrity of the tissue is not affected by the birth process, and both the fetal vasculature of the placenta and the intervillous space filled with maternal blood remain intact. The structural integrity of the tissue, as well as the fact that metabolic functions of the pla-

form colonies. c HBCs after phagocytosis of fluorescence-­labeled LDL cholesterol. (With kind permission of Jelena Lögl)

centa are still maintained after delivery, provides the basis for studies that can be performed without risk to mother and newborn. In a large number of placental perfusion studies over the past decades, both nutrient and gas exchange between mother and child have been investigated, but also the transport or transfer of antibodies, hormones, growth factors, active substances, nanomaterials or environmental toxins. Placental perfusion also offers the possibility to investigate the function of the pla-

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centa in different pregnancy pathologies, such as preeclampsia, intrauterine growth restriction or gestational diabetes, and to compare them with uncomplicated pregnancies. 15.5.1 

15

Methodology

As mentioned above, the maintenance of placental cell barriers is crucial for the function of the placenta and thus represents a particular challenge for the successful performance of a perfusion experiment. Today, a widely used method is the perfusion of one functional unit of the placenta, a so-called cotyledon. This method is founded on work by Panigel, Schneider and Dancis (Panigel 1962; Schneider et al. 1972) and has been continuously developed. In addition to tissue integrity, an important factor is the shortest possible time interval between placental delivery and cannulation of the corresponding fetal vascular pair (artery and vein) in order to keep the ischemic period as short as possible. Initially, fetal blood is flushed out via the cannulated artery and exits the placenta via the cannulated fetal vein. Subsequently, the selected cotyledon can be fixed in a temperature-controlled perfusion chamber, connected to a tube and pump system and supplied with culture medium. To ensure the most physiological conditions possible, the culture medium is heated to 37°C and the oxygen content is reduced in the fetal circulation using a gas mixture of N2 and CO2. The integrity of the fetal vasculature is checked at this time by the volume of circulating culture medium. An intact vascular system returns >95% of the medium used to the vein exiting the placenta. The next step is to establish maternal circulation, this is done by passing blunt cannulas through the basal plate into the intervillous space and pumping the culture medium through it. The culture medium leaves the intervillous space by diffusion and can be returned to the maternal reservoir. After a preperfusion period of about 30 min,

the planned experiment can be performed. Crucial for the success and validity of an ex vivo perfusion experiment are, on the one hand, the guarantee of conditions that are as physiological as possible and, on the other hand, the constant monitoring of parameters that reflect the integrity and vitality of the tissue. The culture medium used should correspond as closely as possible to the composition of maternal and fetal blood. For this purpose, a bicarbonate-buffered basic medium with glucose, amino acids, vitamins and minerals is often mixed with serum albumin (of bovine or human origin) or dextran as a plasma expander. Dissolved gases and thus the degree of oxygenation of the culture medium represent another important class of substances. In blood, hemoglobin is the most important O2 and CO2 transport system. In the placental intervillous space oxygen partial pressures of about 100  mmHg are reached, these are much lower in the umbilical cord blood with 20–30 mmHg. In the perfusion experiment, the culture medium is gassed with a gas mixture of N2 and 5% CO2 in order to approximate the oxygen partial pressure in the fetal medium to physiological values on the one hand and to keep the pH of the bicarbonate buffer system stable at pH 7.4 on the other. The flow rates in the experimental maternal and fetal circulation are adjusted to the average size of a perfused cotyledon to keep the back pressure of the fetal vasculature low and to maintain the integrity of the endothelium. The intervillous space does not have a vascular system, but maternal blood circulates around the fetal villous tissue. To maintain a pressure gradient from the maternal to the fetal circulation, maternal flow rates must be selected approximately three times higher than fetal flow rates. Fetal vascular backpressure is continuously monitored by micropressure catheters or similar sensors. During the establishment phase of the perfusion experiment, fetal placental vascular back pressure continuously decreases until a stable value of >The consequences of maternal hyperglycemia in the child are the effects of fetal hyperinsulinism. If maternal hyperglycemia is avoided, the consequences in the child can be completely avoided.

16.3 

Diagnosis and Therapy

16.3.1 

Screening and Diagnosis of Gestational Diabetes

An evaluation for the presence of risk factors for diabetes should be performed dur-

ing the initial presentation of the pregnant woman. If risk factors are present, a diagnostic 75-g oGTT should be performed as soon as the pregnancy is established. The German maternity guideline prescribes a 50-g screening test for all pregnant women at 24–28  weeks of gestation. Abnormal results must then be confirmed by diagnostic 75-g oGTT (. Fig.  16.2) (AWMF 2011). The cut-­off values of the 75-g-oGTT recommended by the International Association of Diabetes and Pregnancy Study Groups (IADPSG) are shown in the following overview.  

24/0-27+6 50-g test

< 135 mg/dl (7.5 mmol/l)

All values normal

≥ 135 < 200 mg/dl ( 7.5–11.1 mmol/l)

≥ 200 mg/dl (11.1 mmol/l)

75-g-oGTT ≥ 92/180/155 mg/dl (5.1; 10.0; 8.6 mmol/l)

GDM No oGTT

≥1 pathological value

Fasting ≥ 126 mg/dl (6.9 mmol/l) and/or 2 hours ≥ 200 mg/dl (11.1 mmol/l)

16 Exclusion GDM

GDM

Type 2 diabetes diagnosed in pregnancy Determining HbA1c Care as for type 1 or type 2

..      Fig. 16.2  Diagnostic procedure to exclude gestational diabetes (GDM) in pregnancy. (According to the revised version of the German “AWMF-Leitlinie Gestationsdiabetes 2011”, not yet published)

16

353 Maternal Disease Affecting the Placenta: Diabetes Mellitus

Threshold Values of the Oral Glucose Tolerance Test with 75  g Glucose (75-g-oGTT) (IADPSG Criteria) 55 Fasting glucose: ≥5.1  mmol/l (≥92 mg/dl) 55 Glucose after 1  h: ≥10.0  mmol/l (≥180 mg/dl) 55 Glucose after 2  h: ≥8.5  mmol/l (≥153 mg/dl)

16.3.2 

Therapy

The therapy of diabetes in pregnancy aims at maintaining maternal normoglycemia. The available studies on the relationship between maternal blood glucose levels and the fetal consequences of diabetes show a linear relationship and do not allow cut-off values to be defined (Hyperglycemia and Adverse Pregnancy Outcome [HAPO] Study Cooperative Research Group et  al. 2008). Therefore, the setting targets are not yet uniform in the various international guidelines. For Germany, the guidelines for the treatment of gestational diabetes and for the treatment of diabetics in pregnancy recommend a target range that should be interpreted more narrowly or more broadly depending on fetal growth (. Table 16.1) (AWMF 2011; 2014). In order to achieve these adjustment goals, insulin therapy is continuously and closely monitored during pregnancy in the case of pre-existing DM. Existing oral medication for type 2 DM must be switched to insulin as soon as pregnancy is established and ideally even as soon as pregnancy is considered. In the case of GDM, treatment consists of a multi-pillar concept of medical education, self-monitoring of blood glucose, moderate dietary changes, exercise therapy and, if necessary, drug therapy. The education about the recommendations for weight  

..      Table 16.1  Target values for blood glucose control in pregnancy Time

Plasma equivalent mmol/l mg/dl

Fasting

65–95

3.6–5.3

1 h postprandial